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1.
PLoS Negl Trop Dis ; 14(7): e0008534, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32735561

RESUMO

Mast cells are innate effector cells that due to their localization in the tissue form the first line of defense against parasites. We have previously shown that specifically mucosal mast cells were essential for the termination of the intestinal Strongyloides ratti infection. Here, we analyze the impact of mast cells on the immune response and defense against the tissue-dwelling filarial nematode Litomosoides sigmodontis using mast cell-deficient Cpa3cre mice. Despite an increase and an activation of mast cells at the site of infection in wildtype BALB/c mice the outcome of L. sigmodontis infection was not changed in mast cell-deficient BALB/c Cpa3cre mice. In Cpa3cre mice neither vascular permeability induced by blood-sucking mites nor the migration of L3 was altered compared to Cpa3 wildtype littermates. Worm burden in the thoracic cavity was alike in the presence and absence of mast cells during the entire course of infection. Although microfilaremiae in the peripheral blood increased in mast cell-deficient mice at some time points, the infection was cleared with comparable kinetics in the presence and absence of mast cells. Moreover, mast cell deficiency had no impact on the cytokine and antibody response to L. sigmodontis. In summary, our findings suggest that mast cells are not mandatory for the initiation of an appropriate immune response and host defense during L. sigmodontis infection in mice.


Assuntos
Filariose/imunologia , Filarioidea/imunologia , Mastócitos/fisiologia , Animais , Permeabilidade Capilar , Carboxipeptidases A/genética , Carboxipeptidases A/metabolismo , Filariose/parasitologia , Camundongos , Camundongos Endogâmicos BALB C , Infestações por Ácaros , Mutação
2.
PLoS Negl Trop Dis ; 14(7): e0008427, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32628671

RESUMO

A major impediment to eliminate lymphatic filariasis and onchocerciasis is the lack of effective short-course macrofilaricidal drugs or regimens that are proven to be safe for both infections. In this study we tested oxfendazole, an anthelmintic shown to be well tolerated in phase 1 clinical trials. In vitro, oxfendazole exhibited modest to marginal motility inhibition of adult worms of Onchocerca gutturosa, pre-adult worms of Onchocerca volvulus and Onchocerca lienalis microfilariae. In vivo, five days of oral treatments provided sterile cure with up to 100% macrofilaricidal efficacy in the murine Litomosoides sigmodontis model of filariasis. In addition, 10 days of oral treatments with oxfendazole inhibited filarial embryogenesis in patent L. sigmodontis-infected jirds and subsequently led to a protracted but complete clearance of microfilaremia. The macrofilaricidal effect observed in vivo was selective, as treatment with oxfendazole of microfilariae-injected naïve mice was ineffective. Based on pharmacokinetic analysis, the driver of efficacy is the maintenance of a minimal efficacious concentration of approximately 100 ng/ml (based on subcutaneous treatment at 25 mg/kg in mice). From animal models, the human efficacious dose is predicted to range from 1.5 to 4.1 mg/kg. Such a dose has already been proven to be safe in phase 1 clinical trials. Oxfendazole therefore has potential to be efficacious for treatment of human filariasis without causing adverse reactions due to drug-induced microfilariae killing.


Assuntos
Benzimidazóis/farmacologia , Filariose Linfática/tratamento farmacológico , Filarioidea/efeitos dos fármacos , Animais , Anti-Helmínticos/uso terapêutico , Modelos Animais de Doenças , Filariose Linfática/parasitologia , Feminino , Filarioidea/embriologia , Gerbillinae/parasitologia , Camundongos , Camundongos Endogâmicos BALB C , Microfilárias/efeitos dos fármacos , Onchocerca/efeitos dos fármacos , Onchocerca volvulus/efeitos dos fármacos , Oncocercose/tratamento farmacológico
3.
Parasite Immunol ; 42(7): e12708, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32145033

RESUMO

Litomosoides sigmodontis is the only filarial nematode where the full life cycle, from larval delivery to the skin through to circulating microfilaria, can be completed in immunocompetent laboratory mice. It is thus an invaluable tool for the study of filariasis. It has been used for the study of novel anti-helminthic therapeutics, the development of vaccines against filariasis, the development of immunomodulatory drugs for the treatment of inflammatory disease and the study of basic immune responses to filarial nematodes. This review will focus on the latter and aims to summarize how the L sigmodontis model has advanced our basic understanding of immune responses to helminths, led to major discoveries in macrophage biology and provided new insights into the immunological functions of the pleural cavity. Finally, and most importantly L sigmodontis represents a suitable platform to study how host genotype affects immune responses, with the potential for further discovery in myeloid cell biology and beyond.


Assuntos
Filariose/imunologia , Filarioidea/imunologia , Interações Hospedeiro-Parasita/imunologia , Animais , Anti-Helmínticos/farmacologia , Modelos Animais de Doenças , Feminino , Filariose/tratamento farmacológico , Filariose/prevenção & controle , Genótipo , Interações Hospedeiro-Parasita/genética , Estágios do Ciclo de Vida , Camundongos , Camundongos Endogâmicos BALB C , Microfilárias/imunologia , Células Mieloides/imunologia
4.
PLoS Negl Trop Dis ; 14(2): e0008119, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-32107497

RESUMO

Neutrophils are essentially involved in protective immune responses against invading infective larvae of filarial nematodes. The present study investigated the impact of S100A8/S100A9 on protective immune responses against the rodent filarial nematode Litomosoides sigmodontis. S100A9 forms with S100A8 the heterodimer calprotectin, which is expressed by circulating neutrophils and monocytes and mitigates or amplifies tissue damage as well as inflammation depending on the immune environment. Mice deficient for S100A8/A9 had a significantly reduced worm burden in comparison to wildtype (WT) animals 12 days after infection (dpi) with infective L3 larvae, either by the vector or subcutaneous inoculation, the latter suggesting that circumventing natural immune responses within the epidermis and dermis do not alter the phenotype. Nevertheless, upon intradermal injection of L3 larvae, increased total numbers of neutrophils, eosinophils and macrophages were observed within the skin of S100A8/A9-/- mice. Furthermore, upon infection the bronchoalveolar and thoracic cavity lavage of S100A8/A9-/- mice showed increased concentrations of CXCL-1, CXCL-2, CXCL-5, as well as elastase in comparison to the WT controls. Neutrophils from S100A8/A9-/- mice exhibited an increased in vitro activation and reduced L3 larval motility more effectively in vitro compared to WT neutrophils. The depletion of neutrophils from S100A8/A9-/- mice prior to L. sigmodontis infection until 5dpi abrogated the protective effect and led to an increased worm burden, indicating that neutrophils mediate enhanced protective immune responses against invading L3 larvae in S100A8/A9-/- mice. Interestingly, complete circumvention of protective immune responses in the skin and the lymphatics by intravenous injection of L3 larvae reversed the phenotype and resulted in an increased worm burden in S100A8/A9-/- mice. In summary, our results reveal that lack of S100A8/S100A9 triggers L3-induced inflammatory responses, increasing chemokine levels, granulocyte recruitment as well as neutrophil activation and therefore impairs larval migration and susceptibility for filarial infection.


Assuntos
Calgranulina A/metabolismo , Calgranulina B/metabolismo , Filariose/imunologia , Filarioidea/imunologia , Animais , Regulação da Expressão Gênica , Larva/imunologia , Pulmão/parasitologia , Pulmão/patologia , Camundongos , Camundongos Knockout , Neutrófilos/fisiologia
5.
PLoS Negl Trop Dis ; 14(1): e0007957, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31986143

RESUMO

The elimination of filarial diseases such as onchocerciasis and lymphatic filariasis is hampered by the lack of a macrofilaricidal-adult worm killing-drug. In the present study, we tested the in vivo efficacy of AN11251, a boron-pleuromutilin that targets endosymbiotic Wolbachia bacteria from filarial nematodes and compared its efficacy to doxycycline and rifampicin. Doxycycline and rifampicin were previously shown to deplete Wolbachia endosymbionts leading to a permanent sterilization of the female adult filariae and adult worm death in human clinical studies. Twice-daily oral treatment of Litomosoides sigmodontis-infected mice with 200 mg/kg AN11251 for 10 days achieved a Wolbachia depletion > 99.9% in the adult worms, exceeding the Wolbachia reduction by 10-day treatments with bioequivalent human doses of doxycycline and a similar reduction as high-dose rifampicin (35 mg/kg). Wolbachia reductions of > 99% were also accomplished by 14 days of oral AN11251 at a lower twice-daily dose (50 mg/kg) or once-per-day 200 mg/kg AN11251 treatments. The combinations tested of AN11251 with doxycycline had no clear beneficial impact on Wolbachia depletion, achieving a > 97% Wolbachia reduction with 7 days of treatment. These results indicate that AN11251 is superior to doxycycline and comparable to high-dose rifampicin in the L. sigmodontis mouse model, allowing treatment regimens as short as 10-14 days. Therefore, AN11251 represents a promising pre-clinical candidate that was identified in the L. sigmodontis model, and could be further evaluated and developed as potential clinical candidate for human lymphatic filariasis and onchocerciasis.


Assuntos
Antibacterianos/farmacologia , Diterpenos/farmacologia , Filariose/tratamento farmacológico , Filarioidea/efeitos dos fármacos , Compostos Policíclicos/farmacologia , Wolbachia/efeitos dos fármacos , Animais , Boro , Doxiciclina/farmacologia , Feminino , Filariose/microbiologia , Filarioidea/microbiologia , Camundongos Endogâmicos BALB C , Rifampina/farmacologia , Simbiose
6.
Parasitol Res ; 119(2): 473-481, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31897790

RESUMO

Equine ocular setariasis arising mainly from ectopic infestation of Setaria digitata is a common vision impairing ophthalmic disease in India, and the identification of this filarial nematode is based solely on morphology. However, morphological characters alone are inadequate to detect and differentiate S. digitata from its congeners. The present communication reports the first phylogenetic characterization of equine S. digitata from India based on sequences derived from the mitochondrial cytochrome c oxidase subunit 1 (COI), the mitochondrial small subunit ribosomal DNA (12S rDNA), and the nuclear internal transcribed spacer 2 (ITS2). Three isolates were characterized for each gene, and respective sequences were submitted to NCBI database (MN078131, MN078132, and MN095798). The sequences were also compared with the other related sequences available from PubMed around the globe, and phylogenetic analysis was carried out in conjunction with nucleotide homologies. There was no intraspecific variation among the Indian isolates. The phylogenetic analysis of S. digitata, inferred from these genes, showed that the isolate sequences obtained from different host species created a separate monophyletic clade within the genus Setaria with minor sequence variations revealing similar molecular characteristics of S. digitata isolates throughout the globe. In addition, the studied Indian isolates were found closer to Sri Lankan isolates. The S. digitata and S. labiatopapillosa appeared as sister species.


Assuntos
Oftalmopatias/veterinária , Filarioidea/isolamento & purificação , Doenças dos Cavalos/parasitologia , Cavalos/parasitologia , Setaria (Nematoide)/isolamento & purificação , Setaríase/parasitologia , Animais , DNA Intergênico/genética , DNA Ribossômico/genética , Complexo IV da Cadeia de Transporte de Elétrons/genética , Oftalmopatias/parasitologia , Filarioidea/genética , Índia , Filogenia , Reação em Cadeia da Polimerase/veterinária , RNA Ribossômico/genética , Análise de Sequência de DNA , Setaria (Nematoide)/genética
7.
Parasit Vectors ; 12(1): 580, 2019 Dec 11.
Artigo em Inglês | MEDLINE | ID: mdl-31829219

RESUMO

BACKGROUND: Parafilaria bovicola (Nematoda: Filariidae) causes cutaneous bleedings in bovine species. Flies serve as intermediate hosts. In recent years, reports on bovine parafilariosis have become more frequent, corroborating the necessity of reliable diagnostic interventions especially since no molecular or serological test has been available. We aimed to establish a polymerase chain reaction assay to detect DNA of P. bovicola in flies, skin biopsies and serohemorraghic exudates of bleeding spots. METHODS: PCRs targeting the cytochrome c oxidase subunit 1 (cox1) gene and the internal transcribed spacer region (ITS) of the ribosomal RNA gene cluster were evaluated for their diagnostic sensitivity as well as performance and specificity on biopsy and serohemorrhagic exudate samples from P. bovicola-infected cattle. RESULTS: Using serohemorrhagic exudates (n = 6), biopsies (n = 2) and flies (n = 1), the PCR targeting the cox1 gene resulted in a gel band of almost 700 bp. Cloning, sequencing, and removal of primer sequences yielded a 649-bp fragment of the P. bovicola cox1 gene. The PCR targeting the ITS region showed a band of about 1100 bp. Cloning, sequencing, and removal of primer sequences resulted in a 1083 bp stretch of the P. bovicola ITS region. Testing samples from presumably affected animals, the cox1-PCR resulted in bands with the expected size and they were all confirmed as P. bovicola by sequencing. In contrast, the ITS-PCR proved to be less sensitive and less specific and additionally amplified the ITS region of Musca domestica or buttercup DNA. When analysing for sensitivity, the cox1-PCR yielded visible bands up to 2 ng of genomic DNA, whereas the ITS-PCR produced bands up to 3 ng. In a plasmid dilution series, the minimum number of target DNA copies was 102 for the cox1-PCR and 101 in the ITS-PCR. CONCLUSIONS: The evaluated cox1-PCR enables reliable detection of P. bovicola DNA in skin biopsies and serohemorrhagic exudates. This PCR and, to a limited extent, the ITS-PCR, may help evaluate different therapeutic approaches. Furthermore, the cox1-PCR may be useful for epidemiological studies on the geographical distribution of P. bovicola. Further understanding of the epidemiology of this parasite will help develop and implement effective control strategies.


Assuntos
Doenças dos Bovinos/diagnóstico , DNA Espaçador Ribossômico/genética , Complexo IV da Cadeia de Transporte de Elétrons/genética , Filariose/veterinária , Filarioidea/isolamento & purificação , Técnicas de Diagnóstico Molecular/métodos , Reação em Cadeia da Polimerase/métodos , Animais , Biópsia , Bovinos , Doenças dos Bovinos/parasitologia , Exsudatos e Transudatos/parasitologia , Filariose/diagnóstico , Filariose/parasitologia , Filarioidea/enzimologia , Filarioidea/genética , Sensibilidade e Especificidade , Pele/parasitologia
8.
PLoS Negl Trop Dis ; 13(12): e0007908, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31815932

RESUMO

T cell-intrinsic regulation, such as anergy, adaptive tolerance and exhaustion, is central to immune regulation. In contrast to Type 1 and Type 17 settings, knowledge of the intrinsic fate and function of Th2 cells in chronic Type 2 immune responses is lacking. We previously showed that Th2 cells develop a PD-1/PD-L2-dependent intrinsically hypo-responsive phenotype during infection with the filarial nematode Litomosoides sigmodontis, denoted by impaired functionality and parasite killing. This study aimed to elucidate the transcriptional changes underlying Th2 cell-intrinsic hypo-responsiveness, and whether it represents a unique and stable state of Th2 cell differentiation. We demonstrated that intrinsically hypo-responsive Th2 cells isolated from L. sigmodontis infected mice stably retained their dysfunctional Th2 phenotype upon transfer to naïve recipients, and had a divergent transcriptional profile to classical Th2 cells isolated prior to hypo-responsiveness and from mice exposed to acute Type 2 stimuli. Hypo-responsive Th2 cells displayed a distinct transcriptional profile to exhausted CD4+ T cells, but upregulated Blimp-1 and the anergy/regulatory-associated transcription factors Egr2 and c-Maf, and shared characteristics with tolerised T cells. Hypo-responsive Th2 cells increased mRNA expression of the soluble regulatory factors Fgl2, Cd38, Spp1, Areg, Metrnl, Lgals3, and Csf1, and a subset developed a T-bet+IFN-γ+ Th2/Th1 hybrid phenotype, indicating that they were not functionally inert. Contrasting with their lost ability to produce Th2 cytokines, hypo-responsive Th2 cells gained IL-21 production and IL-21R blockade enhanced resistance to L. sigmodontis. IL-21R blockade also increased the proportion of CD19+PNA+ germinal centre B cells and serum levels of parasite specific IgG1. This indicates a novel regulatory role for IL-21 during filarial infection, both in controlling protection and B cell responses. Thus, Th2 cell-intrinsic hypo-responsiveness is a distinct and stable state of Th2 cell differentiation associated with a switch from a classically active IL-4+IL-5+ Th2 phenotype, to a non-classical dysfunctional and potentially regulatory IL-21+Egr2+c-Maf+Blimp-1+IL-4loIL-5loT-bet+IFN-γ+ Th2 phenotype. This divergence towards alternate Th2 phenotypes during chronicity has broad implications for the outcomes and treatment of chronic Type 2-related infections and diseases.


Assuntos
Antígenos de Helmintos/imunologia , Filariose/patologia , Filarioidea/imunologia , Células Th2/imunologia , Animais , Modelos Animais de Doenças , Feminino , Filariose/imunologia , Perfilação da Expressão Gênica , Camundongos Endogâmicos BALB C , Fenótipo , Células Th2/patologia
9.
PLoS Negl Trop Dis ; 13(11): e0007811, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31770367

RESUMO

BACKGROUND: The release of small non-coding RNAs (sRNAs) has been reported in parasitic nematodes, trematodes and cestodes of medical and veterinary importance. However, little is known regarding the diversity and composition of sRNAs released by different lifecycle stages and the portion of sRNAs that persist in host tissues during filarial infection. This information is relevant to understanding potential roles of sRNAs in parasite-to-host communication, as well as to inform on the location within the host and time point at which they can be detected. METHODOLOGY AND PRINCIPAL FINDINGS: We have used small RNA (sRNA) sequencing analysis to identify sRNAs in replicate samples of the excretory-secretory (ES) products of developmental stages of the filarial nematode Litomosoides sigmodontis in vitro and compare this to the parasite-derived sRNA detected in host tissues. We show that all L. sigmodontis developmental stages release RNAs in vitro, including ribosomal RNA fragments, 5'-derived tRNA fragments (5'-tRFs) and, to a lesser extent, microRNAs (miRNAs). The gravid adult females (gAF) produce the largest diversity and abundance of miRNAs in the ES compared to the adult males or microfilariae. Analysis of sRNAs detected in serum and macrophages from infected animals reveals that parasite miRNAs are preferentially detected in vivo, compared to their low levels in the ES products, and identifies miR-92-3p and miR-71-5p as L. sigmodontis miRNAs that are stably detected in host cells in vivo. CONCLUSIONS: Our results suggest that gravid adult female worms secrete the largest diversity of extracellular sRNAs compared to adult males or microfilariae. We further show differences in the parasite sRNA biotype distribution detected in vitro versus in vivo. We identify macrophages as one reservoir for parasite sRNA during infection, and confirm the presence of parasite miRNAs and tRNAs in host serum during patent infection.


Assuntos
Filariose/genética , Filarioidea/genética , Filarioidea/fisiologia , Interações Hospedeiro-Parasita/fisiologia , Pequeno RNA não Traduzido/sangue , Animais , Líquidos Corporais , Feminino , Filariose/parasitologia , Estágios do Ciclo de Vida , Macrófagos , Masculino , Camundongos , MicroRNAs/genética , Microfilárias , RNA Ribossômico , RNA de Transferência , Análise de Sequência
10.
PLoS Negl Trop Dis ; 13(8): e0007636, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31381563

RESUMO

Depletion of Wolbachia endosymbionts of human pathogenic filariae using 4-6 weeks of doxycycline treatment can lead to permanent sterilization and adult filarial death. We investigated the anti-Wolbachia drug candidate ABBV-4083 in the Litomosoides sigmodontis rodent model to determine Wolbachia depletion kinetics with different regimens. Wolbachia reduction occurred in mice as early as 3 days after the initiation of ABBV-4083 treatment and continued throughout a 10-day treatment period. Importantly, Wolbachia levels continued to decline after a 5-day-treatment from 91.5% to 99.9% during a 3-week washout period. In jirds, two weeks of ABBV-4083 treatment (100mg/kg once-per-day) caused a >99.9% Wolbachia depletion in female adult worms, and the kinetics of Wolbachia depletion were recapitulated in peripheral blood microfilariae. Similar to Wolbachia depletion, inhibition of embryogenesis was time-dependent in ABBV-4083-treated jirds, leading to a complete lack of late embryonic stages (stretched microfilariae) and lack of peripheral microfilariae in 5/6 ABBV-4083-treated jirds by 14 weeks after treatment. Twice daily treatment in comparison to once daily treatment with ABBV-4083 did not significantly improve Wolbachia depletion. Moreover, up to 4 nonconsecutive daily treatments within a 14-dose regimen did not significantly erode Wolbachia depletion. Within the limitations of an animal model that does not fully recapitulate human filarial disease, our studies suggest that Wolbachia depletion should be assessed clinically no earlier than 3-4 weeks after the end of treatment, and that Wolbachia depletion in microfilariae may be a viable surrogate marker for the depletion within adult worms. Furthermore, strict daily adherence to the dosing regimen with anti-Wolbachia candidates may not be required, provided that the full regimen is subsequently completed.


Assuntos
Antibacterianos/farmacologia , Filarioidea/microbiologia , Microfilárias/microbiologia , Wolbachia/efeitos dos fármacos , Wolbachia/fisiologia , Animais , Doxiciclina/farmacologia , Feminino , Filariose , Filarioidea/efeitos dos fármacos , Gerbillinae , Cinética , Camundongos , Camundongos Endogâmicos BALB C , Microfilárias/efeitos dos fármacos , Microfilárias/embriologia , Modelos Animais
11.
PLoS Negl Trop Dis ; 13(8): e0007691, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31469835

RESUMO

Lung disease is regularly reported in human filarial infections but the molecular pathogenesis of pulmonary filariasis is poorly understood. We used Litomosoides sigmodontis, a rodent filaria residing in the pleural cavity responsible for pleural inflammation, to model responses to human filarial infections and probe the mechanisms. Wild-type and Th2-deficient mice (ΔdblGata1 and Il-4receptor(r)a-/-/IL-5-/-) were infected with L. sigmodontis. Survival and growth of adult filariae and prevalence and density of microfilariae were evaluated. Cells and cytokines in the pleural cavity and bronchoalveolar space were characterized by imaging, flow cytometry and ELISA. Inflammatory pathways were evaluated by transcriptomic microarrays and lungs were isolated and analyzed for histopathological signatures. 40% of WT mice were amicrofilaremic whereas almost all mutant mice display blood microfilaremia. Microfilariae induced pleural, bronchoalveolar and lung-tissue inflammation associated with an increase in bronchoalveolar eosinophils and perivascular macrophages, production of mucus, visceral pleura alterations and fibrosis. Inflammation and pathology were decreased in Th2-deficient mice. An IL-4R-dependent increase of CD169 was observed on pleural and bronchoalveolar macrophages in microfilaremic mice. CD169+ tissue-resident macrophages were identified in the lungs with specific localizations. Strikingly, CD169+ macrophages increased significantly in the perivascular area in microfilaremic mice. These data describe lung inflammation and pathology in chronic filariasis and emphasize the role of Th2 responses according to the presence of microfilariae. It is also the first report implicating CD169+ lung macrophages in response to a Nematode infection.


Assuntos
Filariose/patologia , Filarioidea/imunologia , Inflamação/patologia , Pulmão/imunologia , Macrófagos/imunologia , Receptores de Interleucina-4/metabolismo , Lectina 1 Semelhante a Ig de Ligação ao Ácido Siálico/análise , Animais , Modelos Animais de Doenças , Feminino , Filariose/imunologia , Inflamação/imunologia , Pulmão/patologia , Macrófagos/química , Camundongos Endogâmicos BALB C , Células Th2/imunologia
12.
Nat Commun ; 10(1): 2895, 2019 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-31263185

RESUMO

Filariases are diseases caused by arthropod-borne filaria nematodes. The related pathologies depend on the location of the infective larvae when their migration, the asymptomatic and least studied phase of the disease, comes to an end. To determine factors assisting in filariae dissemination, we image Litomosoides sigmodontis infective larvae during their escape from the skin. Burrowing through the dermis filariae exclusively enter pre-collecting lymphatics by mechanical disruption of their wall. Once inside collectors, their rapid and unidirectional movement towards the lymph node is supported by the morphology of lymphatic valves. In a microfluidic maze mimicking lymphatic vessels, filariae follow the direction of the flow, the first biomechanical factor capable of helminth guidance within the host. Finally, non-infective nematodes that rely on universal morpho-physiological cues alone also migrate through the dermis, and break in lymphatics, indicating that the ability to spread by the lymphatic route is an ancestral trait rather than acquired parasitic adaptation.


Assuntos
Filariose/parasitologia , Filarioidea/fisiologia , Vasos Linfáticos/parasitologia , Animais , Feminino , Humanos , Sistema Linfático/irrigação sanguínea , Sistema Linfático/parasitologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Pele/parasitologia
13.
Parasitol Res ; 118(7): 2097-2105, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31154526

RESUMO

In Germany, knowledge of disease agents transmitted by arthropods in zoological gardens is scarce. In the framework of ecological studies, mosquitoes were therefore collected in German zoological gardens and examined for mosquito-borne pathogen DNA and RNA. In total, 3840 mosquitoes were screened for filarial nematodes and three groups of viruses (orthobunyaviruses, flaviviruses, alphaviruses) while 405 mosquitoes were tested for avian malaria parasites. In addition to the filarial nematode species Dirofilaria repens (n = 1) and Setaria tundra (n = 8), Sindbis virus (n = 1) and the haemosporidian genera Haemoproteus (n = 8), Leucocytozoon (n = 10) and Plasmodium (n = 1) were demonstrated. Identified pathogens have the potential to cause disease in zoo and wild animals, but some of them also in humans. Positive mosquitoes were collected most often in July, indicating the highest infection risk during this month. Most of the pathogens were found in mosquito specimens of the Culex pipiens complex, suggesting that its members possibly act as the most important vectors in the surveyed zoos, although the mere demonstration of pathogen DNA/RNA in a homogenised complete mosquito is not finally indicative for a vector role. Outcomes of the study are not only significant for arthropod management in zoological gardens, but also for the general understanding of the occurrence and spread of mosquito-borne disease agents.


Assuntos
Culicidae/parasitologia , Filarioidea/classificação , Haemosporida/classificação , Malária Aviária/parasitologia , Mosquitos Vetores/parasitologia , Plasmodium/classificação , Animais , Culex/parasitologia , Feminino , Filarioidea/genética , Filarioidea/isolamento & purificação , Jardins , Alemanha/epidemiologia , Haemosporida/genética , Haemosporida/isolamento & purificação , Humanos , Malária Aviária/epidemiologia , Malária Aviária/transmissão , Plasmodium/genética , Plasmodium/isolamento & purificação
14.
Parasit Vectors ; 12(1): 248, 2019 May 20.
Artigo em Inglês | MEDLINE | ID: mdl-31109364

RESUMO

BACKGROUND: Mice are susceptible to infections with the rodent filarial nematode Litomosoides sigmodontis and develop immune responses that resemble those of human filarial infections. Thus, the L. sigmodontis model is used to study filarial immunomodulation, protective immune responses against filariae and to screen drug candidates for human filarial diseases. While previous studies showed that type 2 immune responses are protective against L. sigmodontis, the present study directly compared the impact of eosinophils, IL-5, and the IL-4R on the outcome of L. sigmodontis infection. METHODS: Susceptible wildtype (WT) BALB/c mice, BALB/c mice lacking eosinophils (dblGATA mice), IL-5-/- mice, IL-4R-/- mice and IL-4R-/-/IL-5-/- mice were infected with L. sigmodontis. Analyses were performed during the peak of microfilaremia in WT animals (71 dpi) as well as after IL-4R-/-/IL-5-/- mice showed a decline in microfilaremia (119 dpi) and included adult worm counts, peripheral blood microfilariae levels, cytokine production from thoracic cavity lavage, the site of adult worm residence, and quantification of major immune cell types within the thoracic cavity and spleen. RESULTS: Our study reveals that thoracic cavity eosinophil numbers correlated negatively with the adult worm burden, whereas correlations of alternatively activated macrophage (AAM) numbers with the adult worm burden (positive correlation) were likely attributed to the accompanied changes in eosinophil numbers. IL-4R-/-/IL-5-/- mice exhibited an enhanced embryogenesis achieving the highest microfilaremia with all animals becoming microfilariae positive and had an increased adult worm burden combined with a prolonged adult worm survival. CONCLUSIONS: These data indicate that mice deficient for IL-4R-/-/IL-5-/- have the highest susceptibility for L. sigmodontis infection, which resulted in an earlier onset of microfilaremia, development of microfilaremia in all animals with highest microfilariae loads, and an extended adult worm survival.


Assuntos
Suscetibilidade a Doenças/imunologia , Eosinófilos/imunologia , Filariose/imunologia , Interleucina-5/genética , Receptores de Superfície Celular/genética , Animais , Modelos Animais de Doenças , Filariose/sangue , Filarioidea/fisiologia , Técnicas de Inativação de Genes , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Microfilárias/imunologia , Ácaros/parasitologia , Transdução de Sinais , Baço/imunologia
15.
Int J Biol Macromol ; 133: 202-213, 2019 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-30991069

RESUMO

Setaria equina heat shock protein (SeqHSP) 70 gene was characterized, cloned and expressed to recombinant protein (rSeqHSP70). The protein was tested for binding with an anti-filarial drug "diethylcarbamazine citrate (DEC)" by equilibrium dialysis method. Molecular docking was also used to determine the binding sites and residues of binding with DEC. The mice were immunized with the protein alone or bound to DEC. Serum IFN-γ levels in the immunized group with protein-drug complex were significantly higher (P < 0.05) than the protein-immunized group. Mouse anti-SeqHSP70 polyclonal IgG recognized 2 bands at 70 and 75 kDa in S. equina adult worm and human cancer cell lines (HepG2 and MCF-7) extracts. The proliferation assay for mice splenocytes revealed a potentiation and down-regulating effects in non-immunized and immunized groups, respectively with the drug-protein complex. The proliferation and IFN-γ assays for purified human NK cells indicated a potentiating effect of the drug-protein complex (DEC concentration is 50 µM) comparable to the protein. DEC at lower concentration (25 mM) could also show a significant increase (P < 0.05) in IFN-γ. From the results, DEC was postulated to induce conformational changes in the protein exposing more epitopes for NK cell binding and activation.


Assuntos
Dietilcarbamazina/metabolismo , Filarioidea/genética , Proteínas de Choque Térmico HSP70/genética , Proteínas de Choque Térmico HSP70/metabolismo , Proteínas de Helminto/genética , Proteínas de Helminto/metabolismo , Adjuvantes Imunológicos/química , Adjuvantes Imunológicos/metabolismo , Adjuvantes Imunológicos/farmacologia , Sequência de Aminoácidos , Animais , Sequência de Bases , Proliferação de Células/efeitos dos fármacos , Clonagem Molecular , Reações Cruzadas , Expressão Gênica , Proteínas de Choque Térmico HSP70/química , Proteínas de Choque Térmico HSP70/farmacologia , Proteínas de Helminto/química , Proteínas de Helminto/farmacologia , Humanos , Interferon gama/metabolismo , Células Matadoras Naturais/citologia , Células Matadoras Naturais/efeitos dos fármacos , Camundongos , Simulação de Acoplamento Molecular , Conformação Proteica , Análise de Sequência , Baço/imunologia
16.
Microbiol Spectr ; 7(2)2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30953430

RESUMO

The Wolbachia endosymbionts encompass a large group of intracellular bacteria of biomedical and veterinary relevance, closely related to Anaplasma, Ehrlichia, and Rickettsia. This genus of Gram-negative members of the Alphaproteobacteria does not infect vertebrates but is instead restricted to ecdysozoan species, including terrestrial arthropods and a family of parasitic filarial nematodes, the Onchocercidae. The Wolbachia profoundly impact not only the ecology and evolution but also the reproductive biology of their hosts, through a wide range of symbiotic interactions. Because they are essential to the survival and reproduction of their filarial nematode hosts, they represent an attractive target to fight filariasis. Their abilities to spread through insect populations and to affect vector competence through pathogen protection have made Wolbachia a staple for controlling vector-borne diseases. Estimated to be present in up to 66% of insect species, the Wolbachia are probably the most abundant endosymbionts on earth. Their success resides in their unique capacity to infect and manipulate the host germ line to favor their vertical transmission through the maternal lineage. Because the Wolbachia resist genetic manipulation and growth in axenic culture, our understanding of their biology is still in its infancy. Despite these limitations, the "-omics" revolution combined with the use of well-established and emerging experimental host models is accelerating our comprehension of the host phenotypes caused by Wolbachia, and the identification of Wolbachia effectors is ongoing.


Assuntos
Interações entre Hospedeiro e Microrganismos/fisiologia , Wolbachia/fisiologia , Animais , Artrópodes/microbiologia , Filarioidea/microbiologia , Insetos/microbiologia , Simbiose , Wolbachia/classificação , Wolbachia/genética , Wolbachia/patogenicidade
17.
PLoS Negl Trop Dis ; 13(2): e0007159, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30818326

RESUMO

There is a significant need for improved treatments for onchocerciasis and lymphatic filariasis, diseases caused by filarial worm infection. In particular, an agent able to selectively kill adult worms (macrofilaricide) would be expected to substantially augment the benefits of mass drug administration (MDA) with current microfilaricides, and to provide a solution to treatment of onchocerciasis / loiasis co-infection, where MDA is restricted. We have identified a novel macrofilaricidal agent, Tylosin A (TylA), which acts by targeting the worm-symbiont Wolbachia bacterium. Chemical modification of TylA leads to improvements in anti-Wolbachia activity and oral pharmacokinetic properties; an optimized analog (ABBV-4083) has been selected for clinical evaluation.


Assuntos
Antibacterianos/farmacologia , Descoberta de Drogas , Filaricidas/farmacologia , Tilosina/análogos & derivados , Tilosina/farmacologia , Wolbachia/efeitos dos fármacos , Animais , Antibacterianos/farmacocinética , Filariose Linfática/tratamento farmacológico , Feminino , Filaricidas/farmacocinética , Filarioidea/efeitos dos fármacos , Filarioidea/microbiologia , Gerbillinae , Camundongos , Camundongos Endogâmicos BALB C , Oncocercose/tratamento farmacológico , Simbiose/efeitos dos fármacos
18.
FASEB J ; 33(5): 6497-6513, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30807258

RESUMO

A type 1 immune response is involved in atherosclerosis progression, whereas the role of a type 2 polarization, especially with regard to an enhanced T helper (Th)2 cell differentiation, is still unclear. Helminths trigger type 2 immune responses, protecting the host from inflammatory disorders. We investigated whether an increased type 2 polarization by administration of Litomosoides sigmodontis adult worm extract (LsAg) affects atherosclerosis in apolipoprotein E-deficient (ApoE-/-) mice. Injections of 50 µg LsAg, i.p. into ApoE-/- mice induced a type 2 immune response shown by increased frequencies of peritoneal eosinophils and alternatively activated macrophages. To analyze the effect of LsAg on atherosclerosis initiation, ApoE-/- mice received a high-fat diet for 12 wk and weekly injections of 50 µg LsAg from wk 5 to 12. Therapeutic effects on advanced atherosclerosis were analyzed in mice that were fed a high-fat diet for 12 wk followed by 12 wk of normal chow and weekly LsAg injections. Both preventive and therapeutic LsAg application significantly decreased plaque size. Therapeutic treatment even caused regression of plaque size and macrophage density in the aortic root and reduced Th1-specific gene expression and intraplaque inflammation. In addition, plaque size after therapeutic treatment was inversely correlated with plaque-infiltrated alternatively activated macrophages. In vitro, LsAg treatment of HUVECs reduced intracellular levels of phosphorylated NF-κB-p65, IκB-α, and JNK1/2. In bifurcation flow-through slides, THP-1 cell adhesion to a HUVEC monolayer was decreased by LsAg in regions of nonuniform shear stress. Applying inhibitors of the respective kinases suggests JNK1/2 inhibition is involved in the suppressed cell adhesion. A switch to an enhanced type 2 immune response by LsAg exerts antiatherogenic effects on murine plaque development, indicating a protective role of a hampered type 1 polarization. In vitro, LsAg affects endothelial signaling pathways, among which JNK1/2 inhibition seems to be involved in the suppression of monocytic cell adhesion under proatherogenic shear stress.-Constanze, K., Tauchi, M., Furtmair, R., Urschel, K., Raaz-Schrauder, D., Neumann, A.-L., Frohberger, S. J., Hoerauf, A., Regus, S., Lang, W., Sagban, T. A., Stumpfe, F. M., Achenbach, S., Hübner, M. P., Dietel, B. Filarial extract of Litomosoides sigmodontis induces a type 2 immune response and attenuates plaque development in hyperlipidemic ApoE-knockout mice.


Assuntos
Aterosclerose/tratamento farmacológico , Misturas Complexas , Filarioidea/química , Hiperlipidemias/tratamento farmacológico , Placa Aterosclerótica/tratamento farmacológico , Células Th2/imunologia , Animais , Aterosclerose/induzido quimicamente , Aterosclerose/genética , Aterosclerose/imunologia , Misturas Complexas/química , Misturas Complexas/farmacologia , Gorduras na Dieta/efeitos adversos , Gorduras na Dieta/farmacologia , Hiperlipidemias/induzido quimicamente , Hiperlipidemias/genética , Hiperlipidemias/imunologia , Camundongos , Camundongos Knockout para ApoE , Placa Aterosclerótica/induzido quimicamente , Placa Aterosclerótica/imunologia , Células Th1/imunologia , Células Th1/patologia
19.
Parasitol Res ; 118(2): 539-549, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30643971

RESUMO

Worldwide approximately 68 million people are infected with lymphatic filariasis (Lf), provoked by Wuchereria bancrofti, Brugia malayi and Brugia timori. This disease can lead to massive swelling of the limbs (elephantiasis) and disfigurement of the male genitalia (hydrocele). Filarial induced immune regulation is characterised by dominant type 2 helper T cell and regulatory immune responses. In vitro studies have provided evidence that signalling via Toll-like receptor-mediated pathways is triggered by filarial associated factors. Nevertheless, until now, less is known about the role of the adapter molecule TRIF during in vivo infections. Here, we used the rodent-specific nematode Litomosoides sigmodontis to investigate the role of TLR signalling and the corresponding downstream adapter and regulatory molecules TRIF, MyD88, IRF1 and IRF3 during an ongoing infection in semi-susceptible C57BL/6 mice. Interestingly, lack of the central adapter molecule TRIF led to higher worm burden and reduced overall absolute cell numbers in the thoracic cavity (the site of infection) 30 days post-infection. In addition, frequencies of macrophages and lymphocytes in the TC were increased in infected TRIF-/- C57BL/6 mice, whereas frequencies of eosinophils, CD4+ and CD8+ T cells were reduced. Nevertheless, cytokine levels and regulatory T cell populations remained comparable between TRIF-deficient and wildtype C57BL/6 mice upon 30 days of L. sigmodontis infection. In summary, this study revealed a crucial role of the adapter molecule TRIF on worm recovery and immune cell recruitment into the site of infection 30 days upon L. sigmodontis infection in C57BL/6 mice.


Assuntos
Proteínas Adaptadoras de Transporte Vesicular/metabolismo , Filariose/imunologia , Filariose/parasitologia , Filarioidea/crescimento & desenvolvimento , Filarioidea/imunologia , Transdução de Sinais , Proteínas Adaptadoras de Transporte Vesicular/genética , Animais , Citocinas/imunologia , Macrófagos/imunologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Linfócitos T Reguladores/imunologia , Células Th2/imunologia
20.
PLoS Negl Trop Dis ; 13(1): e0006320, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30650105

RESUMO

Flubendazole (FBZ) is highly efficacious against filarial nematodes after parenteral administration and presents a promising macrofilaricidal drug candidate for the elimination of onchocerciasis and other filariae. In the present study the efficacy of a newly developed bioavailable amorphous solid dispersion (ASD) oral formulation of FBZ was investigated in the Litomosoides sigmodontis jird model. FBZ was administered to chronically infected, microfilariae-positive jirds by single (40mg/kg), repeated (2, 6 or 15mg/kg for 5 or 10 days) oral (OR) doses or single subcutaneous (SC) injections (2 or 10mg/kg). Jirds treated with 5 SC injections at 10mg/kg served as positive controls, with untreated animals used as negative controls. After OR doses, FBZ is rapidly absorbed and cleared and the exposures increased dose proportionally. SC administered FBZ was slowly released from the injection site and plasma levels remained constant up to necropsy eight weeks after treatment end. Increasing single SC doses caused less than dose-proportional exposures. At necropsy, all animals receiving 1x or 5x 10mg/kg SC FBZ had cleared all adult worms and the 1x 2mg/kg SC treatment had reduced the adult worm burden by 98%. 10x 15mg/kg OR FBZ reduced the adult worm burden by 95%, whereas 1x 40mg/kg and 5x 15mg/kg OR reduced the worm burden by 85 and 84%, respectively. Microfilaremia was completely cleared at necropsy in all animals of the SC treatment regimens, while all oral FBZ treatment regimens reduced the microfilaremia by >90% in a dose and duration dependent manner. In accordance, embryograms from female worms revealed a FBZ dose and duration dependent inhibition of embryogenesis. Histological analysis of the remaining female adult worms showed that FBZ had damaged the body wall, intestine and most prominently the uterus and uterine content. Results of this study demonstrate that single and repeated SC injections and repeated oral administrations of FBZ have an excellent macrofilaricidal effect.


Assuntos
Filariose/tratamento farmacológico , Filaricidas/farmacologia , Filaricidas/farmacocinética , Filarioidea/efeitos dos fármacos , Mebendazol/análogos & derivados , Administração Oral , Animais , Modelos Animais de Doenças , Desenvolvimento Embrionário/efeitos dos fármacos , Feminino , Filariose/parasitologia , Filaricidas/administração & dosagem , Filarioidea/embriologia , Gerbillinae/parasitologia , Mebendazol/administração & dosagem , Mebendazol/farmacocinética , Mebendazol/farmacologia , Carga Parasitária
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