RESUMO
Ascariasis is a global health problem for humans and animals. Adult Ascaris nematodes are long-lived in the host intestine where they interact with host cells as well as members of the microbiota resulting in chronic infections. Nematode interactions with host cells and the microbial environment are prominently mediated by parasite-secreted proteins and peptides possessing immunomodulatory and antimicrobial activities. Previously, we discovered the C-type lectin protein AsCTL-42 in the secreted products of adult Ascaris worms. Here we tested recombinant AsCTL-42 for its ability to interact with bacterial and host cells. We found that AsCTL-42 lacks bactericidal activity but neutralized bacterial cells without killing them. Treatment of bacterial cells with AsCTL-42 reduced invasion of intestinal epithelial cells by Salmonella. Furthermore, AsCTL-42 interacted with host myeloid C-type lectin receptors. Thus, AsCTL-42 is a parasite protein involved in the triad relationship between Ascaris, host cells, and the microbiota.
Assuntos
Ascaris suum/metabolismo , Interações Hospedeiro-Parasita , Mucosa Intestinal/metabolismo , Lectinas Tipo C/metabolismo , Lectinas/metabolismo , Salmonella , Animais , Ascaríase/metabolismo , Ascaríase/microbiologia , Ascaris suum/microbiologia , Ascaris suum/fisiologia , Linhagem Celular , Lectinas/fisiologia , Proteínas Recombinantes , Sus scrofa/microbiologia , Sus scrofa/parasitologiaRESUMO
Ascaris spp. undergo extensive migration within the body before establishing patent infections in the small intestinal tract of humans and pigs. However, whether larval migration is critical for inducing efficient type 2 responses remains poorly understood. Therefore, we investigated systemic versus local adaptive immune responses along the hepato-tracheal migration of Ascaris suum during primary, single infections in conventionally raised pigs. Neither the initial invasion of gut tissue nor migration through the liver resulted in discernable Th2 cell responses. In contrast, lung-stage larvae elicited a Th2-biased pulmonary response, which declined after the larvae had left the lungs. In the small intestine, we observed an accumulation of Th2 cells upon the arrival of fourth-stage larvae (L4) to the small intestinal lumen. In parallel, we noticed robust and increasing Th1 responses in circulation, migration-affected organs, and draining lymph nodes. Phenotypic analysis of CD4+ T cells specifically recognizing A. suum antigens in the circulation and lung tissue of infected pigs confirmed that the majority of Ascaris-specific T cells produced IL-4 (Th2) and, to a much lesser extent, IL-4/IFN-g (Th2/1 hybrids) or IFN-g alone (Th1). These data demonstrate that lung-stage but not the early liver-stage larvae lead to a locally restricted Th2 response. Significant Th2 cell accumulation in the small intestine occurs only when L4 complete the body migration. In addition, Th2 immunity seems to be hampered by the concurrent, nonspecific Th1 bias in growing pigs. Together, the late onset of Th2 immunity at the site of infection and the Th1-biased systemic immunity likely enable the establishment of intestinal infections by sufficiently large L4 stages and pre-adult worms, some of which resist expulsion mechanisms.
Assuntos
Ascaríase , Ascaris suum , Células Th1 , Células Th2 , Animais , Ascaris suum/imunologia , Ascaríase/imunologia , Ascaríase/parasitologia , Células Th2/imunologia , Suínos , Células Th1/imunologia , Doenças dos Suínos/imunologia , Doenças dos Suínos/parasitologia , Pulmão/imunologia , Pulmão/parasitologia , Larva/imunologia , Citocinas/metabolismoRESUMO
Natural killer (NK) cells play a key role in defense against Salmonella infections during the early phase of infection. Our previous work showed that the excretory/secretory products of Ascaris suum repressed NK activity in vitro. Here, we asked if NK cell functionality was influenced in domestic pigs during coinfection with Ascaris and Salmonella enterica serotype Typhimurium. Ascaris coinfection completely abolished the IL-12 and IL-18 driven elevation of IFN-γ production seen in CD16 + CD8α + perforin + NK cells of Salmonella single-infected pigs. Furthermore, Ascaris coinfection prohibited the Salmonella-driven rise in NK perforin levels and CD107a surface expression. In line with impaired effector functions, NK cells from Ascaris-single and coinfected pigs displayed elevated expression of the inhibitory KLRA1 and NKG2A receptors genes, contrasting with the higher expression of the activating NKp46 and NKp30 receptors in NK cells during Salmonella single infection. These differences were accompanied by the highly significant upregulation of T-bet protein expression in NK cells from Ascaris-single and Ascaris/Salmonella coinfected pigs. Together, our data strongly indicate a profound repression of NK functionality by an Ascaris infection which may hinder infected individuals from adequately responding to a concurrent bacterial infection.
Assuntos
Ascaríase , Coinfecção , Células Matadoras Naturais , Doenças dos Suínos , Animais , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/metabolismo , Ascaríase/imunologia , Ascaríase/veterinária , Ascaríase/parasitologia , Coinfecção/imunologia , Coinfecção/microbiologia , Coinfecção/parasitologia , Suínos , Doenças dos Suínos/parasitologia , Doenças dos Suínos/imunologia , Doenças dos Suínos/microbiologia , Salmonelose Animal/imunologia , Salmonella typhimurium/imunologia , Salmonella typhimurium/patogenicidade , Ascaris suum/imunologia , Interferon gama/metabolismo , Perforina/metabolismo , Interleucina-12/metabolismo , Proteínas com Domínio T/metabolismo , Proteínas com Domínio T/genética , Interleucina-18/metabolismoRESUMO
Ascaris is one of the most widespread helminth infections, leading to chronic morbidity in humans and considerable economic losses in pig farming. In addition, pigs are an important reservoir for the zoonotic salmonellosis, where pigs can serve as asymptomatic carriers. Here, we investigated the impact of an ongoing Ascaris infection on the immune response to Salmonella in pigs. We observed higher bacterial burdens in experimentally coinfected pigs compared to pigs infected with Salmonella alone. The impaired control of Salmonella in the coinfected pigs was associated with repressed interferon gamma responses in the small intestine and with the alternative activation of gut macrophages evident in elevated CD206 expression. Ascaris single and coinfection were associated with a rise of CD4-CD8α+FoxP3+ Treg in the lymph nodes draining the small intestine and liver. In addition, macrophages from coinfected pigs showed enhanced susceptibility to Salmonella infection in vitro and the Salmonella-induced monocytosis and tumor necrosis factor alpha production by myeloid cells was repressed in pigs coinfected with Ascaris. Hence, our data indicate that acute Ascaris infection modulates different immune effector functions with important consequences for the control of tissue-invasive coinfecting pathogens.IMPORTANCEIn experimentally infected pigs, we show that an ongoing infection with the parasitic worm Ascaris suum modulates host immunity, and coinfected pigs have higher Salmonella burdens compared to pigs infected with Salmonella alone. Both infections are widespread in pig production and the prevalence of Salmonella is high in endemic regions of human Ascariasis, indicating that this is a clinically meaningful coinfection. We observed the type 2/regulatory immune response to be induced during an Ascaris infection correlates with increased susceptibility of pigs to the concurrent bacterial infection.
RESUMO
The ascarids are a large group of parasitic nematodes that infect a wide range of animal species. In humans, they cause neglected diseases of poverty; many animal parasites also cause zoonotic infections in people. Control measures include hygiene and anthelmintic treatments, but they are not always appropriate or effective and this creates a continuing need to search for better ways to reduce the human, welfare and economic costs of these infections. To this end, Le Studium Institute of Advanced Studies organized a two-day conference to identify major gaps in our understanding of ascarid parasites with a view to setting research priorities that would allow for improved control. The participants identified several key areas for future focus, comprising of advances in genomic analysis and the use of model organisms, especially Caenorhabditis elegans, a more thorough appreciation of the complexity of host-parasite (and parasite-parasite) communications, a search for novel anthelmintic drugs and the development of effective vaccines. The participants agreed to try and maintain informal links in the future that could form the basis for collaborative projects, and to co-operate to organize future meetings and workshops to promote ascarid research.
Assuntos
Anti-Helmínticos , Zoonoses , Animais , Humanos , Zoonoses/prevenção & controle , Caenorhabditis elegans , Academias e Institutos , Pesquisa , Anti-Helmínticos/uso terapêuticoRESUMO
BACKGROUND: Intestinal helminths are extremely prevalent among humans and animals. In particular, intestinal roundworms affect more than 1 billion people around the globe and are a major issue in animal husbandry. These pathogens live in intimate contact with the host gut microbiota and harbor bacteria within their own intestines. Knowledge of the bacterial host microbiome at the site of infection is limited, and data on the parasite microbiome is, to the best of our knowledge, non-existent. RESULTS: The intestinal microbiome of the natural parasite and zoonotic macropathogen, Ascaris suum was analyzed in contrast to the diversity and composition of the infected host gut. 16S sequencing of the parasite intestine and host intestinal compartments showed that the parasite gut has a significantly less diverse microbiome than its host, and the host gut exhibits a reduced microbiome diversity at the site of parasite infection in the jejunum. While the host's microbiome composition at the site of infection significantly determines the microbiome composition of its parasite, microbial signatures differentiate the nematodes from their hosts as the Ascaris intestine supports the growth of microbes that are otherwise under-represented in the host gut. CONCLUSION: Our data clearly indicate that a nematode infection reduces the microbiome diversity of the host gut, and that the nematode gut represents a selective bacterial niche harboring bacteria that are derived but distinct from the host gut. Video Abstract.
Assuntos
Ascaris suum , Microbioma Gastrointestinal , Helmintos , Microbiota , Nematoides , Parasitos , Humanos , Animais , Bactérias/genéticaRESUMO
Ascariasis is a globally spread intestinal nematode infection of humans and a considerable concern in pig husbandry. Ascaris accomplishes a complex body migration from the intestine via the liver and lung before returning to the intestine. Tissue migration and the habitat shared with a complex microbial community pose the question of how the nematode interacts with microbes and host cells from various tissues. This review addresses the current knowledge of the trilateral relationship between Ascaris, its microbial environment, and host cells, and discusses novel approaches targeting these interactions to combat this widespread infection of livestock and man.
Assuntos
Ascaríase/veterinária , Microbioma Gastrointestinal/fisiologia , Interações Hospedeiro-Parasita , Doenças dos Suínos/microbiologia , Doenças dos Suínos/parasitologia , Animais , Ascaríase/microbiologia , Ascaríase/parasitologia , Ascaris/fisiologia , Meio Ambiente , SuínosRESUMO
Ascaris spp. is a major health problem of humans and animals alike, and understanding the immunogenicity of its antigens is required for developing urgently needed vaccines. The parasite-secreted products represent the most relevant, yet complex (>250 proteins) antigens of Ascaris spp. as defining the pathogen-host interplay. We applied an in vitro antigen processing system coupled to quantitative proteomics to identify potential CD4+ Th cell epitopes in Ascaris-secreted products. This approach considerably restricts the theoretical list of epitopes using conventional CD4+ Th cell epitope prediction tools. We demonstrate the specificity and utility of our approach on two sets of candidate lists, allowing us identifying hits excluded by either one or both computational methods. More importantly, one of the candidates identified experimentally, clearly demonstrates the presence of pathogen-reactive T cells in healthy human individuals against these antigens. Thus, our work pipeline identifies the first human T cell epitope against Ascaris spp. and represents an easily adaptable platform for characterization of complex antigens, in particular for those pathogens that are not easily amenable for in vivo experimental validation.
RESUMO
Aberrant cholesterol metabolism is increasingly appreciated to be essential for prostate cancer initiation and progression. Transcript expression of the high-density lipoprotein-cholesterol receptor scavenger receptor B1 (SR-B1) is elevated in primary prostate cancer. Hypothesizing that SR-B1 expression may help facilitate malignant transformation, we document increased SR-B1 protein and transcript expression in prostate cancer relative to normal prostate epithelium that persists in lethal castration-resistant prostate cancer (CRPC) metastasis. As intratumoral steroid synthesis from the precursor cholesterol can drive androgen receptor (AR) pathway activity in CRPC, we screened androgenic benign and cancer cell lines for sensitivity to SR-B1 antagonism. Benign cells were insensitive to SR-B1 antagonism, and cancer line sensitivity inversely correlated with expression levels of full-length and splice variant AR. In androgen-responsive CRPC cell model C4-2, SR-B1 antagonism suppressed cholesterol uptake, de novo steroidogenesis, and AR activity. SR-B1 antagonism also suppressed growth and viability and induced endoplasmic reticulum stress and autophagy. The inability of exogenous steroids to reverse these effects indicates that AR pathway activation is insufficient to overcome cytotoxic stress caused by a decrease in the availability of cholesterol. Furthermore, SR-B1 antagonism decreased cholesterol uptake, growth, and viability of the AR-null CRPC cell model PC-3, and the small-molecule SR-B1 antagonist block lipid transport-1 decreased xenograft growth rate despite poor pharmacologic properties. Overall, our findings show that SR-B1 is upregulated in primary and castration-resistant disease and is essential for cholesterol uptake needed to drive both steroidogenic and nonsteroidogenic biogenic pathways, thus implicating SR-B1 as a novel and potentially actionable target in CRPC. SIGNIFICANCE: These findings highlight SR-B1 as a potential target in primary and castration-resistant prostate cancer that is essential for cholesterol uptake needed to drive steroidogenic and nonsteroidogenic biogenic pathways.
Assuntos
Androgênios/metabolismo , Neoplasias Ósseas/secundário , Colesterol/metabolismo , Neoplasias Hepáticas/secundário , Neoplasias Pulmonares/secundário , Neoplasias de Próstata Resistentes à Castração/patologia , Receptores Depuradores Classe B/metabolismo , Animais , Apoptose , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Neoplasias Ósseas/metabolismo , Neoplasias Ósseas/cirurgia , Proliferação de Células , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/cirurgia , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/cirurgia , Masculino , Camundongos , Camundongos Nus , Orquiectomia , Prognóstico , Neoplasias de Próstata Resistentes à Castração/metabolismo , Neoplasias de Próstata Resistentes à Castração/cirurgia , Receptores Androgênicos/genética , Receptores Androgênicos/metabolismo , Receptores Depuradores Classe B/genética , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Gastrointestinal nematodes are among the most prevalent parasites infecting humans and livestock worldwide. Infective larvae of the soil-transmitted nematode Ascaris spp. enter the host and start tissue migration by crossing the intestinal epithelial barrier. The initial interaction of the intestinal epithelium with the parasite, however, has received little attention. In a time-resolved interaction model of porcine intestinal epithelial cells (IPEC-J2) and infective Ascaris suum larvae, we addressed the early transcriptional changes occurring simultaneously in both organisms using dual-species RNA-Seq. Functional analysis of the host response revealed an overall induction of metabolic activity, without induction of immune responsive genes or immune signaling pathways and showing suppression of chemotactic genes like CXCL8/IL-8 or CHI3L1. Ascaris larvae, when getting in contact with the epithelium, showed induction of genes that orchestrate motor activity and larval development, such as myosin, troponin, myoglobin, and protein disulfide isomerase 2 (PDI-2). In addition, excretory-secretory products that likely facilitate parasite invasion were increased, among them, aspartic protease 6 or hyaluronidase. Integration of host and pathogen data in an interspecies gene co-expression network indicated links between nematode fatty acid biosynthesis and host ribosome assembly/protein synthesis. In summary, our study provides new molecular insights into the early factors of parasite invasion, while at the same time revealing host immunological unresponsiveness. Reproducible software for dual RNA-Seq analysis of non-model organisms is available at https://gitlab.com/mkuhring/project_asuum and can be applied to similar studies.
Assuntos
Regulação da Expressão Gênica , Helmintos/imunologia , Enteropatias Parasitárias/genética , Enteropatias Parasitárias/imunologia , Mucosa Intestinal/imunologia , Mucosa Intestinal/metabolismo , Transcriptoma , Animais , Biologia Computacional/métodos , Perfilação da Expressão Gênica , Ontologia Genética , Interações Hospedeiro-Parasita/genética , Interações Hospedeiro-Parasita/imunologia , Enteropatias Parasitárias/parasitologia , Mucosa Intestinal/parasitologia , Larva , SuínosRESUMO
[This corrects the article DOI: 10.3389/fcimb.2018.00271.].
RESUMO
Ascariasis is a widespread soil-transmitted helminth infection caused by the intestinal roundworm Ascaris lumbricoides in humans, and the closely related Ascaris suum in pigs. Progress has been made in understanding interactions between helminths and host immune cells, but less is known concerning the interactions of parasitic nematodes and the host microbiota. As the host microbiota represents the direct environment for intestinal helminths and thus a considerable challenge, we studied nematode products, including excretory-secretory products (ESP) and body fluid (BF), of A. suum to determine their antimicrobial activities. Antimicrobial activities against gram-positive and gram-negative bacterial strains were assessed by the radial diffusion assay, while effects on biofilm formation were assessed using the crystal violet static biofilm and macrocolony assays. In addition, bacterial neutralizing activity was studied by an agglutination assay. ESP from different A. suum life stages (in vitro-hatched L3, lung-stage L3, L4, and adult) as well as BF from adult males were analyzed by mass spectrometry. Several proteins and peptides with known and predicted roles in nematode immune defense were detected in ESP and BF samples, including members of A. suum antibacterial factors (ASABF) and cecropin antimicrobial peptide families, glycosyl hydrolase enzymes such as lysozyme, as well as c-type lectin domain-containing proteins. Native, unconcentrated nematode products from intestine-dwelling L4-stage larvae and adults displayed broad-spectrum antibacterial activity. Additionally, adult A. suum ESP interfered with biofilm formation by Escherichia coli, and caused bacterial agglutination. These results indicate that A. suum uses a variety of factors with broad-spectrum antibacterial activity to affirm itself within its microbe-rich environment in the gut.
Assuntos
Antibacterianos/metabolismo , Antibiose , Ascaris suum/metabolismo , Bactérias/efeitos dos fármacos , Bactérias/crescimento & desenvolvimento , Biofilmes/efeitos dos fármacos , Biofilmes/crescimento & desenvolvimento , Testes de Aglutinação , Animais , Antibacterianos/análise , Ascaris suum/química , Violeta Genciana/análise , Proteínas de Helminto/análise , Proteínas de Helminto/metabolismo , Espectrometria de Massas , Testes de Sensibilidade Microbiana , Coloração e Rotulagem , SuínosRESUMO
Intestinal parasitic nematodes live in intimate contact with the host microbiota. Changes in the microbiome composition during nematode infection affect immune control of the parasites and shifts in the abundance of bacterial groups have been linked to the immunoregulatory potential of nematodes. Here we asked if the small intestinal parasite Heligmosomoides polygyrus produces factors with antimicrobial activity, senses its microbial environment and if the anti-nematode immune and regulatory responses are altered in mice devoid of gut microbes. We found that H. polygyrus excretory/secretory products exhibited antimicrobial activity against gram+/- bacteria. Parasites from germ-free mice displayed alterations in gene expression, comprising factors with putative antimicrobial functions such as chitinase and lysozyme. Infected germ-free mice developed increased small intestinal Th2 responses coinciding with a reduction in local Foxp3+RORγt+ regulatory T cells and decreased parasite fecundity. Our data suggest that nematodes sense their microbial surrounding and have evolved factors that limit the outgrowth of certain microbes. Moreover, the parasites benefit from microbiota-driven immune regulatory circuits, as an increased ratio of intestinal Th2 effector to regulatory T cells coincides with reduced parasite fitness in germ-free mice.
Assuntos
Microbioma Gastrointestinal/imunologia , Enteropatias Parasitárias/imunologia , Nematospiroides dubius/imunologia , Infecções por Strongylida/imunologia , Animais , Bactérias/classificação , Bactérias/genética , Bactérias/imunologia , Análise por Conglomerados , Genes de Helmintos/genética , Interações entre Hospedeiro e Microrganismos/imunologia , Enteropatias Parasitárias/parasitologia , Intestino Delgado/microbiologia , Intestino Delgado/parasitologia , Camundongos Endogâmicos C57BL , Nematospiroides dubius/genética , Nematospiroides dubius/fisiologia , Organismos Livres de Patógenos Específicos , Infecções por Strongylida/parasitologia , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/metabolismo , Células Th2/imunologia , Células Th2/metabolismo , Transcriptoma/imunologiaRESUMO
Parasitic nematode infections are widespread in nature, affecting humans as well as wild, companion, and livestock animals. Most parasitic nematodes inhabit the intestines of their hosts living in close contact with the intestinal microbiota. Many species also have tissue migratory life stages in the absence of severe systemic inflammation of the host. Despite the close coexistence of helminths with numerous microbes, little is known concerning these interactions. While the environmental niche is considerably different, the free-living nematode Caenorhabditis elegans (C. elegans) is also found amongst a diverse microbiota, albeit on decaying organic matter. As a very well characterized model organism that has been intensively studied for several decades, C. elegans interactions with bacteria are much more deeply understood than those of their parasitic counterparts. The enormous breadth of understanding achieved by the C. elegans research community continues to inform many aspects of nematode parasitology. Here, we summarize what is known regarding parasitic nematode-bacterial interactions while comparing and contrasting this with information from work in C. elegans. This review highlights findings concerning responses to bacterial stimuli, antimicrobial peptides, and the reciprocal influences between nematodes and their environmental bacteria. Furthermore, the microbiota of nematodes as well as alterations in the intestinal microbiota of mammalian hosts by helminth infections are discussed.
Assuntos
Microbiologia Ambiental , Microbioma Gastrointestinal/fisiologia , Interações Hospedeiro-Parasita/fisiologia , Intestinos/microbiologia , Nematoides/microbiologia , Animais , Fenômenos Fisiológicos Bacterianos , Biodiversidade , Caenorhabditis elegans/microbiologia , Caenorhabditis elegans/fisiologia , Resistência Microbiana a Medicamentos , Fezes/microbiologia , Microbioma Gastrointestinal/imunologia , Helmintíase/microbiologia , Helmintos/microbiologia , Interações Hospedeiro-Parasita/efeitos dos fármacos , Interações Hospedeiro-Parasita/imunologia , Humanos , Modelos Biológicos , Nematoides/classificação , Nematoides/efeitos dos fármacos , Nematoides/imunologia , Transdução de SinaisRESUMO
Protonated nanostructured aluminum silicate (NSAS) is a protonated montmorillonite clay that was shown to be effective as an inhibitor of intestinal cholesterol absorption. The effect of NSAS on the intestinal absorption of nutrients is unknown. An in vitro lipolysis model was adapted to test the intraluminal processing of vitamin D3 and K1 in the presence of various amounts of NSAS. Additionally, vitamin absorption was assessed in male Sprague-Dawley rats randomized in the following treatment groups: IV administration of 0.1 mg/kg vitamin D3 and 1 mg/kg vitamin K1, and a single-dose gavage of 1 mg/kg vitamin D3 and 5mg/kg of vitamin K1 in peanut oil with various doses of NSAS slurry, 2% NSAS-fortified diet, or 50 mg/kg stigmastanol. The solubilized fraction of vitamin D3 in the lipolysis medium was reduced from 70% to 46% upon the addition of 120 mg NSAS. In contrast, the solubilized fractions of vitamin K1 were not significantly affected. Although the NSAS-fortified diet did not significantly affect the absorbed fraction of both vitamins, NSAS slurry decreased the absorption of vitamin D3 as compared to the control. These results indicate that NSAS may be incorporated in diet to treat hypercholesterolemia; however, vitamin D3 monitoring may be required.