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1.
Exp Parasitol ; 221: 108060, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-33338467

RESUMO

Amoebic keratitis (AK) is a sight-threatening infection characterized by a severe inflammation of the cornea, caused by the free-living protozoan of the genus Acanthamoeba. Identification of amoebic proteins involved in AK pathogenesis may help to elucidate molecular mechanisms of infection and contribute to indicate diagnostic and therapeutic targets. In this study, we evaluated changes in the expression profile of Acanthamoeba proteins triggered by the invasive process, using an approach involving two-dimensional polyacrylamide gel electrophoresis (2DE PAGE), followed by mass spectrometry identification (ESI-IT-TOF LC-MSn). AK was induced by intrastromal inoculation in Wistar rats, using trophozoites from a T4 genotype, human case-derived A. castellanii strain under prolonged axenic culture. Cultures re-isolated from the lesions after two successive passages in the animals were used as biological triplicate for proteomic experiments. Analysis of the protein profile comparing long-term and re-isolated cultures indicated 62 significant spots, from which 27 proteins could be identified in the Acanthamoeba proteome database. Five of them (Serpin, Carboxypeptidase A1, Hypothetical protein, Calponin domain-containing protein, aldo/keto reductase) were exclusively found in the re-isolated trophozoites. Our analysis also revealed that a concerted modulation of several biochemical pathways is triggered when A. castellanii switches from a free-living style to a parasitic mode, including energetic metabolism, proteolytic activity, control of gene expression, protein degradation and methylation of DNA, which may be also involved in gain of virulence in an animal model of AK.


Assuntos
Ceratite por Acanthamoeba/metabolismo , Acanthamoeba castellanii/metabolismo , Proteínas de Protozoários/biossíntese , Ceratite por Acanthamoeba/parasitologia , Análise de Variância , Animais , Modelos Animais de Doenças , Humanos , Masculino , Proteômica , Proteínas de Protozoários/genética , Ratos , Ratos Wistar , Espectrometria de Massas por Ionização por Electrospray , Eletroforese em Gel Diferencial Bidimensional
2.
Exp Parasitol ; 218: 108008, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-32979343

RESUMO

Acanthamoeba sp. is a free living amoeba that causes severe, painful and fatal infections, viz. Acanthamoeba keratitis and granulomatous amoebic encephalitis among humans. Antimicrobial chemotherapy used against Acanthamoeba is toxic to human cells and show side effects as well. Infections due to Acanthamoeba also pose challenges towards currently used antimicrobial treatment including resistance and transformation of trophozoites to resistant cyst forms that can lead to recurrence of infection. Therapeutic agents targeting central nervous system infections caused by Acanthamoeba should be able to cross blood-brain barrier. Nanoparticles based drug delivery put forth an effective therapeutic method to overcome the limitations of currently used antimicrobial chemotherapy. In recent years, various researchers investigated the effectiveness of nanoparticles conjugated drug and/or naturally occurring plant compounds against both trophozoites and cyst form of Acanthamoeba. In the current review, a reasonable effort has been made to provide a comprehensive overview of various nanoparticles tested for their efficacy against Acanthamoeba. This review summarizes the noteworthy details of research performed to elucidate the effect of nanoparticles conjugated drugs against Acanthamoeba.


Assuntos
Acanthamoeba/efeitos dos fármacos , Amebicidas/administração & dosagem , Nanopartículas/administração & dosagem , Acanthamoeba/crescimento & desenvolvimento , Ceratite por Acanthamoeba/tratamento farmacológico , Ceratite por Acanthamoeba/parasitologia , Amebíase/tratamento farmacológico , Amebíase/mortalidade , Amebíase/parasitologia , Amebicidas/farmacologia , Amebicidas/uso terapêutico , Biguanidas/administração & dosagem , Biguanidas/farmacologia , Biguanidas/uso terapêutico , Infecções Protozoárias do Sistema Nervoso Central/tratamento farmacológico , Infecções Protozoárias do Sistema Nervoso Central/mortalidade , Infecções Protozoárias do Sistema Nervoso Central/parasitologia , Clorexidina/administração & dosagem , Clorexidina/farmacologia , Clorexidina/uso terapêutico , Sistemas de Liberação de Medicamentos , Imunocompetência , Hospedeiro Imunocomprometido , Encefalite Infecciosa/tratamento farmacológico , Encefalite Infecciosa/mortalidade , Encefalite Infecciosa/parasitologia , Nanopartículas/classificação , Nanopartículas/uso terapêutico , Trofozoítos/efeitos dos fármacos
3.
Parasitol Res ; 119(6): 1943-1954, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32385711

RESUMO

Acanthamoeba causes diseases such as Acanthamoeba keratitis (AK) which leads to permanent blindness and granulomatous Acanthamoeba encephalitis (GAE) where there is formation of granulomas in the brain. Current treatments such as chlorhexidine, diamidines, and azoles either exhibit undesirable side effects or require immediate and prolonged treatment for the drug to be effective or prevent relapse. Previously, antifungal drugs amphotericin B, nystatin, and fluconazole-conjugated silver with nanoparticles have shown significantly increased activity against Acanthamoeba castellanii. In this study, two functionally diverse tetrazoles were synthesized, namely 5-(3-4-dimethoxyphenyl)-1H-tetrazole and 1-(3-methoxyphenyl)-5-phenoxy-1H-tetrazole, denoted by T1 and T2 respectively. These compounds were evaluated for anti-Acanthamoeba effects at different concentrations ranging from 5 to 50 µM. Furthermore, these compounds were conjugated with silver nanoparticles (AgNPs) to enhance their efficacy. Particle size analysis showed that T1-AgNPs and T2-AgNPs had an average size of 52 and 70 nm respectively. After the successful synthesis and characterization of tetrazoles and tetrazole-conjugated AgNPs, they were subjected to anti-Acanthamoeba studies. Amoebicidal assay showed that at concentration 10 µM and above, T2 showed promising antiamoebic activities between the two compounds while encystation and excystation assays reveal that both T1 and T2 have inhibited differentiation activity against Acanthamoeba castellanii. Conjugation of T1 and T2 to AgNP also increased efficacy of tetrazoles as anti-Acanthamoeba agents. This may be due to the increased bioavailability as AgNP allows better delivery of treatment compounds to A. castellanii. Human cell cytotoxicity assay revealed that tetrazoles and AgNPs are significantly less toxic towards human cells compared with chlorhexidine which is known to cause undesirable side effects. Cytopathogenicity assay also revealed that T2 conjugated with AgNPs significantly reduced cytopathogenicity of A. castellanii compared with T2 alone, suggesting that T2-conjugated AgNP is an effective and safe anti-Acanthamoeba agent. The use of a synthetic azole compound conjugated with AgNPs can be an alternative strategy for drug development against A. castellanii. However, mechanistic and in vivo studies are needed to explore further translational values.


Assuntos
Acanthamoeba castellanii/efeitos dos fármacos , Amebicidas/farmacologia , Nanopartículas Metálicas , Prata/farmacologia , Tetrazóis/farmacologia , Ceratite por Acanthamoeba/tratamento farmacológico , Ceratite por Acanthamoeba/parasitologia , Acanthamoeba castellanii/genética , Acanthamoeba castellanii/isolamento & purificação , Amebicidas/síntese química , Amebicidas/toxicidade , Clorexidina/farmacologia , Genótipo , Células HeLa , Humanos , Tetrazóis/síntese química , Tetrazóis/toxicidade
4.
Exp Parasitol ; 215: 107915, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32461112

RESUMO

Acanthamoeba castellanii is an opportunistic protozoan responsible for serious human infections including Acanthamoeba keratitis and granulomatous amoebic encephalitis. Despite advances in antimicrobial therapy and supportive care, infections due to Acanthamoeba are a major public concern. Current methods of treatment are not fully effective against both the trophozoite and cyst forms of A. castellanii and are often associated with severe adverse effects, host cell cytotoxicity and recurrence of infection. Therefore, there is an urgent need to develop new therapeutic approaches for the treatment and management of Acanthamoebic infections. Repurposing of clinically approved drugs is a viable avenue for exploration and is particularly useful for neglected and rare diseases where there is limited interest by pharmaceutical companies. Nanotechnology-based drug delivery systems offer promising approaches in the biomedical field, particularly in diagnosis and drug delivery. Herein, we conjugated an antihyperglycemic drug, metformin with silver nanoparticles and assessed its anti-acanthamoebic properties. Characterization by ultraviolet-visible spectrophotometry and atomic force microscopy showed successful formation of metformin-coated silver nanoparticles. Amoebicidal and amoebistatic assays revealed that metformin-coated silver nanoparticles reduced the viability and inhibited the growth of A. castellanii significantly more than metformin and silver nanoparticles alone at both 5 and 10 µM after 24 h incubation. Metformin-coated silver nanoparticles also blocked encystation and inhibited the excystation in Acanthamoeba after 72 h incubation. Overall, the conjugation of metformin with silver nanoparticles was found to enhance its antiamoebic effects against A. castellanii. Furthermore, the pretreatment of A. castellanii with metformin and metformin-coated silver nanoparticles for 2 h also reduced the amoebae-mediated host cell cytotoxicity after 24 h incubation from 73% to 10% at 10 µM, indicating that the drug-conjugated silver nanoparticles confer protection to human cells. These findings suggest that metformin-coated silver nanoparticles hold promise in the improved treatment and management of Acanthamoeba infections.


Assuntos
Acanthamoeba castellanii/efeitos dos fármacos , Metformina/administração & dosagem , Ceratite por Acanthamoeba/tratamento farmacológico , Ceratite por Acanthamoeba/parasitologia , Anti-Infecciosos Locais/farmacologia , Infecções Protozoárias do Sistema Nervoso Central/tratamento farmacológico , Infecções Protozoárias do Sistema Nervoso Central/parasitologia , Clorexidina/farmacologia , Células HeLa , Humanos , Encefalite Infecciosa/tratamento farmacológico , Encefalite Infecciosa/parasitologia , Nanopartículas Metálicas/administração & dosagem , Nanopartículas Metálicas/uso terapêutico , Metformina/farmacologia , Metformina/uso terapêutico , Microscopia de Força Atômica , Encistamento de Parasitas/efeitos dos fármacos , Prata , Espectrofotometria Ultravioleta , Trofozoítos/efeitos dos fármacos
5.
PLoS One ; 15(3): e0229681, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32160218

RESUMO

PURPOSE: To determine whether Acanthamoeba keratitis (AK) patients have higher rates of Acanthamoeba and free-living amoeba (FLA) colonising domestic sinks than control contact lens (CL) wearers, and whether these isolates are genetically similar to the corneal isolates from their CL associated AK. METHODS: 129 AK patients from Moorefield Eye Hospital, London and 64 control CL wearers from the Institute of Optometry were included in this study. The participants self-collected home kitchen and bathroom samples from tap-spouts, overflows and drains using an instructional kit. The samples were cultured by inoculating onto a non-nutrient agar plate seeded with Escherichia coli, incubated at 32°C and examined for amoebae by microscopy for up to 2 weeks. Partial sequences of mitochondrial cytochrome oxidase genes (coxA) of Acanthamoeba isolates from four AK patients were compared to Acanthamoeba isolated from the patient's home. The association between sampling sites was analysed with the chi-square test. RESULTS: A total of 513 samples from AK patients and 189 from CL controls were collected. The yield of FLA was significantly greater in patients' bathrooms (72.1%) than CL controls' bathrooms (53.4%) (p<0.05). Spouts (kitchen 6.7%, bathroom 11%) had the lowest rate of Acanthamoeba isolation compared to drains (kitchen 18.2%, bathroom 27.9%) and overflow (kitchen 39.1%, bathroom 25.9%) either in kitchens or bathrooms (p<0.05). There was no statistically significant difference between the average prevalence of Acanthamoeba in all three sample sites in kitchens (16.9%) compared to all three sample sites in bathrooms (21.5%) and no association for Acanthamoeba prevalence between AK patients and CL controls. All four corneal isolates had the same coxA sequence as at least one domestic water isolate from the patients' sink of the kitchen and the bathroom. CONCLUSION: The prevalence of Acanthamoeba and FLA was high in UK homes. FLA colonisation was higher in AK patients compared to controls but the prevalence of Acanthamoeba between AK patients and CL controls domestic sinks was similar. This study confirms that domestic water isolates are probably the source of AK infection. Advice about avoiding water contact when using CL's should be mandatory.


Assuntos
Ceratite por Acanthamoeba/epidemiologia , Ceratite por Acanthamoeba/parasitologia , Acanthamoeba/isolamento & purificação , Acanthamoeba/genética , Acanthamoeba/patogenicidade , Amebozoários/isolamento & purificação , Estudos de Casos e Controles , Lentes de Contato/efeitos adversos , Lentes de Contato/parasitologia , Suscetibilidade a Doenças , Meio Ambiente , Habitação , Humanos , Londres/epidemiologia , Fatores de Risco , Engenharia Sanitária , Água/parasitologia
6.
Parasit Vectors ; 13(1): 123, 2020 Mar 07.
Artigo em Inglês | MEDLINE | ID: mdl-32143731

RESUMO

BACKGROUND: Acanthamoeba spp. are ubiquitous pathogens which cause granulomatous amoebic encephalitis and disseminated infection. Moreover, Acanthamoeba spp. infection of the cornea leads to Acanthamoeba keratitis. Our previous study showed that the infection of an eyeball may also take place via the migration of trophozoites through the optic nerve from the brain to the eyes. The aim of the study was to analyze the activity of enzymatic antioxidants and the concentration of non-enzymatic antioxidant in the eyes of immunocompetent and immunocompromised mice with disseminated acanthamoebiasis. RESULTS: In the immunocompetent mice infected with Acanthamoeba spp. we noted a significant decrease in catalase activity at 8 and 16 days post-infection (dpi). Glutathione reductase activity was significantly lower at 16 dpi compared to the control group and glutathione concentration was statistically higher at 24 dpi than in the control group. In the immunosuppressed mice, a statistically significant increase in glutathione concentration in the eye samples was found at 16 dpi compared to those not infected with Acanthamoeba spp. In the immunosuppressed mice infected with Acanthamoeba spp., glutathione peroxidase activity was statistically lower at 8 dpi, and glutathione concentration was statistically significantly higher at 16 dpi compared to the control group. CONCLUSIONS: The inflammatory response in the eyes of hosts with experimental acanthamoebiasis led to changes in the activity of enzymatic antioxidants and the content of non-enzymatic antioxidant. Therefore, the dysregulation of antioxidants may play a role in the pathomechanism of Acanthamoeba eye infection.


Assuntos
Ceratite por Acanthamoeba/tratamento farmacológico , Ceratite por Acanthamoeba/metabolismo , Acanthamoeba/metabolismo , Antioxidantes/farmacologia , Hospedeiro Imunocomprometido , Acanthamoeba/imunologia , Ceratite por Acanthamoeba/parasitologia , Ceratite por Acanthamoeba/patologia , Animais , Antioxidantes/uso terapêutico , Catalase/metabolismo , Modelos Animais de Doenças , Olho/imunologia , Olho/patologia , Glutationa , Glutationa Peroxidase/metabolismo , Glutationa Redutase/metabolismo , Camundongos , Oxirredução
7.
J Fr Ophtalmol ; 43(4): 330-333, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32151474

RESUMO

Acanthamoeba keratitis due to a genus of free-living amoebae is a severe corneal infection. Treatment of this disease is based on the combined use of antiseptics and other drugs, including azoles. We tested isavuconazole, the latest marketed azole, in vitro, against A. castellanii, A. lenticulata and A. hatchetti. Our results show that isavuconazole presents slight amoebistatic activity against A. castellanii trophozoites but no cysticidal activity. Isavuconazole could be used only in association for management of AK due to A. castellanii.


Assuntos
Ceratite por Acanthamoeba/parasitologia , Acanthamoeba/efeitos dos fármacos , Nitrilas/farmacologia , Piridinas/farmacologia , Triazóis/farmacologia , Acanthamoeba/classificação , Acanthamoeba/crescimento & desenvolvimento , Acanthamoeba/fisiologia , Ceratite por Acanthamoeba/tratamento farmacológico , Acanthamoeba castellanii/efeitos dos fármacos , Acanthamoeba castellanii/crescimento & desenvolvimento , Acanthamoeba castellanii/fisiologia , Animais , Relação Dose-Resposta a Droga , Humanos , Nitrilas/uso terapêutico , Encistamento de Parasitas/efeitos dos fármacos , Testes de Sensibilidade Parasitária , Piridinas/uso terapêutico , Triazóis/uso terapêutico , Trofozoítos/efeitos dos fármacos
8.
J Med Microbiol ; 69(1): 87-95, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31846414

RESUMO

Introduction. Acanthamoeba keratitis is a sight-threatening corneal infection that is commonly reported among contact lens users and those suffering from corneal trauma. The prevalence of Acanthamoeba species or genotypes in causing keratitis infection is not well known.Aim. This study was conducted to identify and genotype Acanthamoeba isolates from keratitis patients, targeting the ribosomal nuclear subunit (Rns) region, and describe the associated clinical presentation and treatment outcome.Methodology. Thirty culture-confirmed patients with Acanthamoeba keratitis, identified in a tertiary eye care centre in South India during the period from December 2016 to December 2018, were included in this study. The data collected from patient records include demographic details, history of illness, mode of trauma, treatment history and follow-up status. The genotype and species were identified based on the Rns sequence and phylogenetic tree analysis.Results. Acanthamoeba culbertsoni was the most predominant keratitis-causing species, followed by Acanthamoeba quina, Acanthamoeba castellanii, Acanthamoeba healyi, Acanthamoeba hatchetti, Acanthamoeba polyphaga and Acanthamoeba stevensoni. Three major genotypes were identified (T4, T11 and T12), with the T4 genotype being the most predominant, with four subclusters, i.e. T4A, T4B, T4D and T4E. This is the first report on corneal infection by the A. stevensoni T11 genotype and the A. healyi T12 genotype. No significant correlation was observed between the clinical outcomes of corneal disease and the genotypes or species.Conclusion. Rns genotyping is very effective in identifying the Acanthamoeba species and genotype in keratitis. Genotyping of Acanthamoeba spp. will help to advance our understanding of genotype-specific pathogenesis and geographical distribution.


Assuntos
Ceratite por Acanthamoeba/patologia , Ceratite por Acanthamoeba/parasitologia , Acanthamoeba castellanii/classificação , Acanthamoeba castellanii/genética , Genótipo , Filogenia , Acanthamoeba castellanii/isolamento & purificação , Genes de RNAr , Técnicas de Genotipagem , Humanos , Índia , Centros de Atenção Terciária
9.
Parasitol Res ; 119(2): 659-666, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31848745

RESUMO

Free-living amoebae of the genus Acanthamoeba are causative agents of Acanthamoeba keratitis and amoebic encephalitis in humans, both of which are serious infections. The ability to produce proteases is one of the factors involved in the pathogenesis of Acanthamoeba infections. The aim of this study was to evaluate the secreted proteases of six Acanthamoeba strains from distinct genotypes (T1, T2, T4 and T11) maintained in prolonged axenic culture and following three successive passages in Madin-Darby Canine Kidney (MDCK) cells. Conditioned medium was obtained from cultures before and after interaction with the MDCK monolayers, resolved in SDS-PAGE containing gelatine, then subjected to quantitative azocasein assays. Zymography profiles varied between the strains, with the predominant proteases found to be serine-type proteases from 49 to 128 kDa. A T1 genotype strain isolated from dust showed quantitatively higher protease secretion compared to the other strains. No changes were detected in the zymography profiles of MDCK-interacted cultures compared to long-term axenic cultures. Two strains presented lower proteolytic activity post-MDCK interaction, while the remaining strains presented similar values before and after MDCK passages. In conclusion, this study confirms the predominance of serine-type protease secretion by Acanthamoeba, with distinct profiles presented by the different strains and genotypes studied. Also, interaction of trophozoites with MDCK cells did not alter the zymography pattern.


Assuntos
Acanthamoeba/enzimologia , Acanthamoeba/metabolismo , Serina Proteases/metabolismo , Acanthamoeba/genética , Ceratite por Acanthamoeba/parasitologia , Animais , Cultura Axênica , Caseínas/análise , Linhagem Celular , Cães , Genótipo , Humanos , Células Madin Darby de Rim Canino , Trofozoítos/metabolismo
10.
Korean J Parasitol ; 57(5): 505-511, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31715691

RESUMO

Acanthamoeba keratitis (AK) is a rare sight-threatening corneal infection, often reporting from contact lens wearers. An asymptomatic human immunodeficiency virus (HIV)-infected Thai male without history of contact lens use complained foreign body sensation at his left eye during motorbike riding. He had neither specific keratitis symptoms nor common drugs responding, which contributed to delayed diagnosis. By corneal re-scraping, Acanthamoeba-like cysts were detected by calcofluor white staining and agar culture. The etiological agent obtained from the culture was molecularly confirmed by Acanthamoeba spp.-specific PCR, followed by DNA sequencing. The results from BLAST and phylogenetic analysis based on the DNA sequences, revealed that the pathogen was Acanthamoeba T4, the major genotype most frequently reported from clinical isolates. The infection was successfully treated with polyhexamethylene biguanide resulting in corneal scar. This appears the first reported AK case from a non-contact lens wearer with HIV infection in Thailand. Although AK is sporadic in developing countries, a role of free-living Acanthamoeba as an opportunistic pathogen should not be neglected. The report would increase awareness of AK, especially in the case presenting unspecific keratitis symptoms without clinical response to empirical antimicrobial therapy.


Assuntos
Ceratite por Acanthamoeba/parasitologia , Acanthamoeba/isolamento & purificação , Infecções por HIV/complicações , Acanthamoeba/classificação , Acanthamoeba/genética , Ceratite por Acanthamoeba/etiologia , Adulto , Doenças Assintomáticas , Olho/parasitologia , Humanos , Masculino , Filogenia , Tailândia
11.
Parasit Vectors ; 12(1): 467, 2019 Oct 09.
Artigo em Inglês | MEDLINE | ID: mdl-31597577

RESUMO

BACKGROUND: Pathogenic protozoans use extracellular vesicles (EVs) for intercellular communication and host manipulation. Acanthamoeba castellanii is a free-living protozoan that may cause severe keratitis and fatal granulomatous encephalitis. Although several secreted molecules have been shown to play crucial roles in the pathogenesis of Acanthamoeba, the functions and components of parasite-derived EVs are far from understood. METHODS: Purified EVs from A. castellanii were confirmed by electron microscopy and nanoparticle tracking analysis. The functional roles of parasite-derived EVs in the cytotoxicity to and immune response of host cells were examined. The protein composition in EVs from A. castellanii was identified and quantified by LC-MS/MS analysis. RESULTS: EVs from A. castellanii fused with rat glioma C6 cells. The parasite-derived EVs induced an immune response from human THP-1 cells and a cytotoxic effect in C6 cells. Quantitative proteomic analysis identified a total of 130 proteins in EVs. Among the identified proteins, hydrolases (50.2%) and oxidoreductases (31.7%) were the largest protein families in EVs. Furthermore, aminopeptidase activities were confirmed in EVs from A. castellanii. CONCLUSIONS: The proteomic profiling and functional characterization of EVs from A. castellanii provide an in-depth understanding of the molecules packaged into EVs and their potential mechanisms mediating the pathogenesis of this parasite.


Assuntos
Acanthamoeba castellanii/fisiologia , Exossomos/química , Exossomos/fisiologia , Proteômica , Ceratite por Acanthamoeba/parasitologia , Acanthamoeba castellanii/patogenicidade , Acanthamoeba castellanii/ultraestrutura , Aminopeptidases/análise , Animais , Infecções Protozoárias do Sistema Nervoso Central/parasitologia , Meios de Cultura , DNA Complementar/biossíntese , Exossomos/imunologia , Exossomos/ultraestrutura , Humanos , Microscopia Eletrônica de Transmissão , Neuroglia/parasitologia , RNA de Protozoário/genética , RNA de Protozoário/isolamento & purificação , Ratos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células THP-1/imunologia , Células THP-1/parasitologia
12.
Indian J Ophthalmol ; 67(11): 1886-1888, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31638063

RESUMO

Identification of Acanthamoeba cysts and trophozoites in cases of keratitis is traditionally done with microbiological techniques such as smear examination with 10% potassium hydroxide (KOH) and culture. Double walled cyst with hexagonal inner wall is characteristic of Acanthamoeba. We report a unique case of a 9 year old boy who presented with dense anterior corneal stromal infltration, which on smear examination showed atypical double walled spherical cysts, leading to a diagnostic dilemma. An 18S rRNA gene-based PCR done on the growth on culture, subsequently identifed a rarely reported species of Acanthamoeba. The patient was advised combination therapy with polyhexamethylene biguanide (PHMB 0.02%) and chlorhexidine (0.02%) eye drops. Three weeks post treatment, the keratitis resolved with scarring and vascularisation and visual acuity improved to 20/60. At 8 weeks follow up Best corrected visual acuity further improved to 20/30 with contact lens.


Assuntos
Ceratite por Acanthamoeba/diagnóstico , Acanthamoeba/genética , Substância Própria/patologia , DNA Ribossômico/análise , Infecções Oculares Parasitárias/diagnóstico , Análise de Sequência de DNA/métodos , Acuidade Visual , Ceratite por Acanthamoeba/parasitologia , Criança , Substância Própria/parasitologia , Infecções Oculares Parasitárias/parasitologia , Humanos , Masculino
13.
Indian J Ophthalmol ; 67(10): 1593-1598, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31546487

RESUMO

Purpose: To understand demographic and socioeconomic barriers and treatment-seeking behaviors of patients with infectious keratitis requiring therapeutic penetrating keratoplasty (TPK) in a developing country. Methods: This prospective non-comparative questionnaire- based study included all patients presenting to Aravind Eye Hospital, Madurai with infectious keratitis that eventuated to TPK between November 2015 and October 2016. A structured questionnaire was administered on post-operative day 3 to collect data on the demographic details, predisposing factors, prior treatment received, and treatment expenditures. Results: In total, 227 patients underwent TPK between November 2015 and October 2016 for infectious keratitis. The majority of patients were males (n = 132, 58.1%), illiterate (n = 129, 56.8%), and had a family monthly income of less than INR 6000 (n = 142, 62.5%). Most of the patients (n = 163, 71.8%) had prior treatment with an ophthalmologist before presenting to our hospital. The mean distance travelled to reach our centre was 269.2 ± 298.5 km. The mean duration of disease before the presentation was 20.3 ± 21.1 days. Corneal smear was positive for fungus in 163 (88.1%) and Aspergillus was the most commonly isolated fungi in 55 (41.3%) cultures. The mean total cost of treatment was INR 8752.87 ± 7615.39 per patient. There was a positive correlation between the duration of the disease (rho 0.19, P = 0.0034) and the costs of treatment (rho 0.2, P = 0.0024) with the distance travelled by the patient. Conclusion: Patients who travelled a farther distance had a delayed onset of presentation and spent significantly more than their respective counterparts.


Assuntos
Ceratite por Acanthamoeba/epidemiologia , Úlcera da Córnea/epidemiologia , Infecções Oculares Bacterianas/epidemiologia , Infecções Oculares Fúngicas/epidemiologia , Ceratoplastia Penetrante , Aceitação pelo Paciente de Cuidados de Saúde/psicologia , Fatores Socioeconômicos , Ceratite por Acanthamoeba/parasitologia , Ceratite por Acanthamoeba/cirurgia , Adulto , Úlcera da Córnea/microbiologia , Úlcera da Córnea/cirurgia , Infecções Oculares Bacterianas/microbiologia , Infecções Oculares Bacterianas/cirurgia , Infecções Oculares Fúngicas/microbiologia , Infecções Oculares Fúngicas/cirurgia , Feminino , Sobrevivência de Enxerto , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Acuidade Visual
14.
Eye Contact Lens ; 45(5): 301-305, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31442212

RESUMO

PURPOSE: To compare the infectious contact lens-related corneal ulcer (CLRU) and non-CLRU cases at Saint Louis University. METHODS: Retrospective review of corneal ulcer cases identified by search of the ophthalmology and microbiology department databases between 1999 and 2016. RESULTS: Six hundred seventy-seven cases of corneal ulcers were identified, of which 46% were CLRU. CLRU cases were seen more commonly in younger patients (P<0.001) and women (P=0.03) than non-CLRU cases. Many of the infections were vision-threatening as defined by central/paracentral location (73% CLRU and 71% non-CLRU [P=0.60]) and large size of ulcer >2 mm in 36% CLRU and 51% non-CLRU (P=0.002). Causative pathogen in cultured CLRU was predominately Pseudomonas species (44%, P<0.001 vs. the non-CLRU group), other gram-negative (6%), gram-positive (33%), fungi (13%), and Acanthamoeba (5%). Comparatively, cultured non-CLRU was predominately gram-positive (64%, P<0.001 vs. the CLRU group), gram-negative (26%), and fungi (11%). The combined oxacillin-resistant Staphylococcus aureus and coagulase-negative Staphylococcus isolates were 35% and 34%, respectively. Despite the progressive increase in the number of corneal ulcers seen, the annual trend for any one particular organism for either CLRU cases or non-CLRU cases did not change significantly. CONCLUSIONS: Most of the cases were non-CLRU. CLRU was disproportionately associated with Pseudomonas species and non-CLRU with Staphylococcal species. Fungal infections were predominately caused by filamentous organisms in both groups. Acanthamoeba keratitis was exclusively associated with CL use.


Assuntos
Ceratite por Acanthamoeba/parasitologia , Lentes de Contato/microbiologia , Lentes de Contato/parasitologia , Úlcera da Córnea/microbiologia , Infecções Oculares Bacterianas/microbiologia , Infecções Oculares Fúngicas/microbiologia , Infecções Oculares Parasitárias/parasitologia , Centros Médicos Acadêmicos , Ceratite por Acanthamoeba/diagnóstico , Ceratite por Acanthamoeba/tratamento farmacológico , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Antibacterianos/uso terapêutico , Antiprotozoários/uso terapêutico , Criança , Pré-Escolar , Úlcera da Córnea/diagnóstico , Úlcera da Córnea/tratamento farmacológico , Infecções Oculares Bacterianas/diagnóstico , Infecções Oculares Bacterianas/tratamento farmacológico , Infecções Oculares Fúngicas/diagnóstico , Infecções Oculares Fúngicas/tratamento farmacológico , Infecções Oculares Parasitárias/diagnóstico , Infecções Oculares Parasitárias/tratamento farmacológico , Feminino , Humanos , Lactente , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos
15.
Korean J Parasitol ; 57(3): 217-223, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31284343

RESUMO

Acanthamoeba castellanii has ubiquitous distribution and causes primary acanthamoebic keratitis (AK). AK is a common disease in contact lens wearers and results in permanent visual impairment or blindness. In this study, we observed the cytopathic effect, in vitro cytotoxicity, and secretion pattern of cytokines in human corneal epithelial cells (HCECs) induced by A. castellanii trophozoites and/or cysts. Morphological observation revealed that panked dendritic HCECs co-cultured with amoeba cysts had changed into round shape and gradually died. Such changes were more severe in co-culture with cyst than those of co-cultivation with trophozoites. In vitro cytotoxicity assay revealed the highest cytotoxicity to HCECs in the co-culture system with amoeba cysts. A. castellanii induced the expression of IL-1α, IL-6, IL-8, and CXCL1 in HCECs. Secreted levels of IL-1α, IL-6, and IL-8 in HCECs co-cultured with both trophozoites and cysts were increased at an early incubation time (3 and 6 hr). These results suggested that cytopathic changes and pro-inflammatory cytokines release of HCECs in response to A. castellanii, especially amoebic cysts, are an important mechanism for AK development.


Assuntos
Ceratite por Acanthamoeba/imunologia , Acanthamoeba castellanii/fisiologia , Córnea/citologia , Células Epiteliais/imunologia , Trofozoítos/fisiologia , Ceratite por Acanthamoeba/parasitologia , Acanthamoeba castellanii/crescimento & desenvolvimento , Células Cultivadas , Córnea/imunologia , Córnea/parasitologia , Células Epiteliais/parasitologia , Humanos , Interleucina-1/genética , Interleucina-1/imunologia , Interleucina-6/genética , Interleucina-6/imunologia , Interleucina-8/genética , Interleucina-8/imunologia , Trofozoítos/crescimento & desenvolvimento
16.
Cornea ; 38(10): 1309-1313, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31306283

RESUMO

PURPOSE: To determine whether combinations of commonly used antiamoebic agents display synergy in their ability to kill Acanthamoeba cysts in vitro. METHODS: Synergy testing was performed with a microdilution checkerboard assay on 10 clinical Acanthamoeba keratitis isolates collected at the Proctor Foundation from 2008 to 2012. Each isolate was exposed to pairwise combinations of chlorhexidine, propamidine, and voriconazole. The minimum cysticidal concentration (MCC) for each drug pair was estimated for each isolate, and the summed fractional cysticidal concentration (ΣFCC) was calculated for each drug combination in the checkerboard, with synergy defined as a lack of growth at a ΣFCC ≤ 0.5 and antagonism as growth at a ΣFCC > 4. RESULTS: Chlorhexidine and propamidine were cysticidal, with median MCCs of 12.5 (range 1.5-50) and 11.7 (range 0.2-250), respectively. Voriconazole was not cysticidal, with a median MCC of >10,000 µg/mL. The combination of chlorhexidine and propamidine did not markedly change the cysticidal activity compared with either drug alone. By contrast, voriconazole antagonized the cysticidal activity of both chlorhexidine and propamidine, with Acanthamoeba growth observed at antagonistic ΣFCCs in 27 of 49 (55.1%, 95% confidence interval 35.7%-78.6%) checkerboard combinations of voriconazole and chlorhexidine and in 58 of 147 (39.5%, 95% confidence interval 14.3%-50.3%) combinations of voriconazole and propamidine. CONCLUSIONS: In an in vitro assay, voriconazole reduced the cysticidal activity of 2 commonly used antiamoebic drugs. Although the in vivo drug interactions could be different, these observations may be useful in cases of nonhealing Acanthamoeba keratitis being treated with combination therapies that include voriconazole.


Assuntos
Ceratite por Acanthamoeba/tratamento farmacológico , Acanthamoeba/isolamento & purificação , Amebicidas/farmacologia , Infecções Oculares Parasitárias/tratamento farmacológico , Voriconazol/farmacologia , Acanthamoeba/efeitos dos fármacos , Ceratite por Acanthamoeba/diagnóstico , Ceratite por Acanthamoeba/parasitologia , Animais , Antifúngicos/farmacologia , Sinergismo Farmacológico , Infecções Oculares Parasitárias/parasitologia , Humanos , Testes de Sensibilidade Parasitária
17.
Parasitol Res ; 118(8): 2443-2454, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31144032

RESUMO

Acanthamoeba keratitis (AK) is a devastating, painful corneal infection, which may lead to loss of vision. The development of resistance and failure of the currently used drugs represent a therapeutic predicament. Thus, novel therapies with lethal effects on resistant Acanthamoeba are necessary to combat AK. In the present study, the curative effect of Nigella sativa aqueous extract (N. sativa) and chitosan nanoparticles (nCs) and both agents combined were assessed in experimentally induced AK. All inoculated corneas developed varying grades of AK. The study medications were applied on the 5th day postinoculation and were evaluated by clinical examination of the cornea and cultivation of corneal scraps. On the 10th day posttreatment, a 100% cure of AK was obtained with nCs (100 µg/ml) in grades 1 and 2 of corneal opacity as well as with N. sativa 60 mg/ml-nCs 100 µg/ml in grades 1, 2, and 3 of corneal opacity, highlighting a possible synergistic effect. On the 15th day posttreatment, a 100% cure was reached with N. sativa aqueous extract (60 mg/ml). Moreover, on the 20th day posttreatment, N. sativa (30 mg/ml) provided a cure rate of 87.5%, while nCs (50 µg/ml) as well as N. sativa 30 mg/ml-nCs 50 µg/ml yielded a cure rate of 75%; the lowest percentage of cure (25%) was obtained with chlorhexidine (0.02%), showing a non-significant difference compared to the parasite control. The clinical outcomes were in agreement with the results of corneal scrap cultivation. The results of the present study demonstrate the effectiveness of N. sativa aqueous extract and nCs (singly or combined) when used against AK, and these agents show potential for the development of new, effective, and safe therapeutic alternatives.


Assuntos
Ceratite por Acanthamoeba/tratamento farmacológico , Antiprotozoários/administração & dosagem , Nigella sativa/química , Extratos Vegetais/administração & dosagem , Acanthamoeba/efeitos dos fármacos , Acanthamoeba/fisiologia , Ceratite por Acanthamoeba/parasitologia , Adulto , Animais , Antiprotozoários/química , Quitosana/química , Quitosana/farmacologia , Quitosana/uso terapêutico , Clorexidina/farmacologia , Córnea/parasitologia , Feminino , Humanos , Masculino , Nanopartículas/química , Extratos Vegetais/química , Ratos , Resultado do Tratamento
18.
Ann Parasitol ; 65(1): 19-25, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31095904

RESUMO

Acanthamoeba keratitis, the vision-threatening corneal disease reported with increasing frequency in Poland is difficult to treat due to extremely high resistance of the amoeba cysts to chemicals. The agents of possible anti-amoebic activity are still tested. Pathogenic Acanthamoeba samples/isolates acquired from severe cases of keratitis examined by molecular techniques to determine genotypes, compared to one another as well as to the environmental Acanthamoeba castellanii Neff strain were included in the studies. These strains were in vitro examined in terms of their sensitivity/resistance to selected chemicals and tolerance to temperature changes. Samples of the strains cultivated in vitro under bacteria-free conditions were monitored during different growth phases. Higher amoebic population dynamics was observed in both pathogenic Acanthamoeba strains during transfer to 37°C. Agents tested influenced population dynamics in different degree; they showed amoebostatic or amoebicidal effects, however a tendency toward induction of encystment also appeared. Because activation of the dormant cysts can lead to repeated development of amoebae, very important is cysticidal efficacy of chemicals. Further in vitro investigations on various Acanthamoeba strains with different chemicals are still necessary.


Assuntos
Acanthamoeba , Amebicidas , Temperatura , Acanthamoeba/efeitos dos fármacos , Acanthamoeba/genética , Ceratite por Acanthamoeba/parasitologia , Amebicidas/farmacologia , Genótipo , Humanos , Polônia , Saúde Pública
19.
Parasitol Res ; 118(7): 2295-2304, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31093751

RESUMO

Acanthamoeba castellanii belonging to the T4 genotype is an opportunistic pathogen which is associated with blinding eye keratitis and rare but fatal central nervous system infection. A. castellanii pose serious challenges in antimicrobial chemotherapy due to its ability to convert into resistant, hardy shell-protected cyst form that leads to infection recurrence. The fatty acid composition of A. castellanii trophozoites is known to be most abundant in oleic acid which chemically is an unsaturated cis-9-Octadecanoic acid and naturally found in animal and vegetable fats and oils. This study was designed to evaluate antiacanthamoebic effects of oleic acid against trophozoites, cysts as well as parasite-mediated host cell cytotoxicity. Moreover, oleic acid-conjugated silver nanoparticles (AgNPs) were also synthesized and tested against A. castellanii. Oleic acid-AgNPs were synthesized by chemical reduction method and characterized by ultraviolet-visible spectrophotometry, atomic force microscopy, dynamic light scattering analysis, and Fourier transform infrared spectroscopy. Viability, growth inhibition, encystation, and excystation assays were performed with 10 and 5 µM concentration of oleic acid alone and oleic acid-conjugated AgNPs. Bioassays revealed that oleic acid alone and oleic acid-conjugated AgNPs exhibited significant antiamoebic properties, whereas nanoparticle conjugation further enhanced the efficacy of oleic acid. Phenotype differentiation assays also showed significant inhibition of encystation and excystation at 5 µM. Furthermore, oleic acid and oleic acid-conjugated AgNPs also inhibited amoebae-mediated host cell cytotoxicity as determined by lactate dehydrogenase release. These findings for the first time suggest that oleic acid-conjugated AgNPs exhibit antiacanthamoebic activity that hold potential for therapeutic applications against A. castellanii.


Assuntos
Ceratite por Acanthamoeba/tratamento farmacológico , Acanthamoeba castellanii/efeitos dos fármacos , Amebicidas/farmacologia , Nanopartículas Metálicas/química , Ácido Oleico/farmacologia , Ceratite por Acanthamoeba/parasitologia , Amebicidas/química , Animais , Olho/parasitologia , Humanos , Microscopia de Força Atômica , Ácido Oleico/química , Prata/química , Espectrofotometria Ultravioleta , Trofozoítos/efeitos dos fármacos
20.
Acta Microbiol Immunol Hung ; 66(3): 289-300, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-30835133

RESUMO

Genus Acanthamoeba is an opportunistic protozoan that is widely distributed in the environment. Within this genus, numerous species are recognized as human pathogens, potentially causing Acanthamoeba keratitis (AK). AK is a corneal disease, associated predominantly with contact lens (CL) wear; its epidemiology is related to the specific Acanthamoeba genotypes. This study reports seven CL wearer, Acanthamoeba PCR-positive patients with AK, diagnosed between January 2015 and 2018. Patients had the diagnosis of AK 1.36 months after first symptoms. Genotyping allowed the identification of six isolates of the T4 and one of the T8 genotypes. At first presentation, pseudendritiformic epithelopathy/dirty epithelium (four eyes, 57.1%), multifocal stromal infiltrates (five eyes, 71.4%), ring infiltrate (three eyes, 42.8%), and perineuritis (one eye, 14.3%) were observed. AK was healed without later recurrence in two eyes (28.5%) using triple-topical therapy, in three eyes (42.8%) following additional penetrating keratoplasty. In one patient (14.3%), AK recurred following successful application of triple-therapy and was treated successfully with repeated triple-topical therapy and in one patient (14.3%), no follow-up data were available after diagnosis. We could not observe correlation of genotype and clinical course or the necessity of corneal transplantation in our case series.


Assuntos
Ceratite por Acanthamoeba/patologia , Ceratite por Acanthamoeba/parasitologia , Acanthamoeba/classificação , Acanthamoeba/genética , Genótipo , Acanthamoeba/isolamento & purificação , Ceratite por Acanthamoeba/terapia , Administração Tópica , Adolescente , Adulto , Antiprotozoários/uso terapêutico , Transplante de Córnea , Feminino , Técnicas de Genotipagem , Humanos , Hungria , Masculino , Pessoa de Meia-Idade , Resultado do Tratamento , Adulto Jovem
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