RESUMEN
No disponible
Asunto(s)
Humanos , Masculino , Anciano , Queratitis/microbiología , Queratitis Herpética/microbiología , Colletotrichum/aislamiento & purificación , Voriconazol/uso terapéutico , Trasplante de CórneaAsunto(s)
Conjuntivitis/microbiología , Dacriocistitis/microbiología , Infecciones Bacterianas del Ojo/microbiología , Queratitis Herpética/microbiología , Staphylococcus aureus Resistente a Meticilina/aislamiento & purificación , Infecciones Estafilocócicas/microbiología , Anciano , Anciano de 80 o más Años , Antibacterianos/uso terapéutico , Conjuntivitis/diagnóstico , Conjuntivitis/tratamiento farmacológico , Estudios Transversales , Dacriocistitis/diagnóstico , Dacriocistitis/tratamiento farmacológico , Dibekacina/análogos & derivados , Dibekacina/uso terapéutico , Infecciones Bacterianas del Ojo/diagnóstico , Infecciones Bacterianas del Ojo/tratamiento farmacológico , Femenino , Humanos , Lactante , Queratitis Herpética/diagnóstico , Queratitis Herpética/tratamiento farmacológico , Masculino , Pruebas de Sensibilidad Microbiana , Infecciones Estafilocócicas/diagnóstico , Infecciones Estafilocócicas/tratamiento farmacológico , Vancomicina/uso terapéuticoRESUMEN
OBJECTIVE: To describe archipelago keratitis, a presumed clinical variant of herpetic epithelial keratitis. DESIGN: Case series. PARTICIPANTS: A series of 6 patients with an unusual form of superficial keratitis. METHODS: History, including age, gender, clinical evolution, and treatment; slit-lamp biomicroscopy findings; in vivo confocal microscopy findings; and corneal epithelial scrapings were analyzed. MAIN OUTCOME MEASURES: Clinical ocular examination, a diagnostic workup including corneal scraping for herpesvirus polymerase chain reaction, in vivo confocal microscopy, and therapeutic outcome. RESULTS: The authors describe a series of 6 patients with keratitis consisting of foci of epithelial erosions associated with subepithelial nummular inflammatory infiltrates and disposed in a radial, centripetal, archipelagolike pattern originating from the limbus. All the patients had a past history of herpetic epithelial keratitis, herpetic vesicles on the ipsilateral lid, or both. Polymerase chain reaction-based screening for herpes simplex virus 1 and 2 in corneal scrapings demonstrated positive results in 2 patients. In vivo corneal confocal microscopy revealed focal areas of hyperreflective epithelial cells and hyperreflective subepithelial dendritic structures overlying activated keratocytes. All the patients improved with oral valacyclovir treatment followed by topical steroid therapy. CONCLUSIONS: Archipelago keratitis may be a new clinical variant of herpetic keratitis, reflecting herpetic dissemination from the limbus to the center of the cornea.
Asunto(s)
Queratitis Herpética/microbiología , Aciclovir/análogos & derivados , Aciclovir/uso terapéutico , Adolescente , Adulto , Anciano , Antivirales/uso terapéutico , ADN Viral/análisis , Quimioterapia Combinada , Epitelio Corneal/microbiología , Herpesvirus Humano 1/genética , Herpesvirus Humano 1/aislamiento & purificación , Herpesvirus Humano 2/genética , Herpesvirus Humano 2/aislamiento & purificación , Humanos , Queratitis Herpética/diagnóstico , Queratitis Herpética/tratamiento farmacológico , Masculino , Microscopía Confocal , Persona de Mediana Edad , Reacción en Cadena de la Polimerasa , Pregnadienos/uso terapéutico , Recurrencia , Valaciclovir , Valina/análogos & derivados , Valina/uso terapéuticoRESUMEN
PURPOSE: To report a case of Paecilomyces lilacinus keratitis, initially misdiagnosed as Penicillium sp., in a patient with a long-standing history of herpes simplex virus (HSV) keratitis. METHODS: A retrospective case report. RESULTS: A 62-year-old man developed P. lilacinus keratitis. He was treated with topical steroids for immune stromal keratitis secondary to HSV before developing the fungal keratitis. Initial corneal cultures were positive for Penicillium sp., but subsequent cultures identified P. lilacinus to be the causative organism. The patient later developed an anterior chamber abscess. Three penetrating keratoplasties, as well as intravitreal injection of amphothericin B, topical miconazole, subconjunctival miconazole, and systemic fluconazole, were required to eradicate the infection. CONCLUSION: To our knowledge, this is a first report of P. lilacinus keratitis in a patient with a previous history of HSV keratitis. The causative organism was initially reported as Penicillium sp. on 2 occasions, before the correct diagnosis was made. Paecilomyces keratitis progressed to an anterior chamber abscess in this eye. Aggressive treatment, including a therapeutic penetrating keratoplasty, intravitreal amphothericin B injection, topical miconazole, and systemic fluconazole can be successful in eradicating this extremely difficult-to-treat infection.
Asunto(s)
Infecciones Fúngicas del Ojo/terapia , Queratitis Herpética/microbiología , Queratitis Herpética/terapia , Micosis/terapia , Paecilomyces/aislamiento & purificación , Anfotericina B/uso terapéutico , Antifúngicos/uso terapéutico , Terapia Combinada , Córnea/microbiología , Quimioterapia Combinada , Infecciones Fúngicas del Ojo/diagnóstico , Infecciones Fúngicas del Ojo/microbiología , Fluconazol/uso terapéutico , Humanos , Queratitis Herpética/diagnóstico , Queratoplastia Penetrante , Masculino , Miconazol/uso terapéutico , Persona de Mediana Edad , Micosis/diagnóstico , Micosis/microbiología , Reoperación , Estudios RetrospectivosRESUMEN
PURPOSE: To report polymicrobial keratitis in a patient with herpetic stromal keratitis. The initial infecting organism, Burkholderia ambifaria, has not previously been reported to cause microbial keratitis. METHODS: Clinical evaluation and corneal culture were performed. RESULTS: A 59-year-old-man undergoing topical corticosteroid therapy for herpes simplex stromal keratitis developed corneal infection with B. ambifaria. The organism was reisolated 12 days after initiation of hourly therapy with topical levofloxacin 0.5%. At reculture Staphylococcus aureus and Enterococcus spp. were also isolated. The addition of topical amikacin and vancomycin led to resolution of the microbial keratitis. CONCLUSIONS: Burkholderia ambifaria infected a compromised cornea, exhibited an unusual sensitivity profile, and remained viable after 12 days of therapy with an antibiotic to which it was sensitive by in vitro tests.
Asunto(s)
Infecciones por Burkholderia , Burkholderia/aislamiento & purificación , Enterococcus/aislamiento & purificación , Infecciones Bacterianas del Ojo , Queratitis Herpética/microbiología , Infecciones Estafilocócicas , Staphylococcus aureus/aislamiento & purificación , Amicacina/uso terapéutico , Infecciones por Burkholderia/diagnóstico , Infecciones por Burkholderia/tratamiento farmacológico , Infecciones por Burkholderia/microbiología , Sustancia Propia/microbiología , Quimioterapia Combinada/uso terapéutico , Infecciones Bacterianas del Ojo/diagnóstico , Infecciones Bacterianas del Ojo/tratamiento farmacológico , Infecciones Bacterianas del Ojo/microbiología , Infecciones por Bacterias Grampositivas/diagnóstico , Infecciones por Bacterias Grampositivas/tratamiento farmacológico , Infecciones por Bacterias Grampositivas/microbiología , Humanos , Queratitis Herpética/diagnóstico , Queratitis Herpética/tratamiento farmacológico , Levofloxacino , Masculino , Persona de Mediana Edad , Ofloxacino/uso terapéutico , Infecciones Estafilocócicas/diagnóstico , Infecciones Estafilocócicas/tratamiento farmacológico , Infecciones Estafilocócicas/microbiología , Vancomicina/uso terapéuticoRESUMEN
PURPOSE: To evaluate the efficacy of amniotic membrane transplantation in the management of treated infectious corneal ulcer in which inflammatory reactions were responsible for corneal damage. METHOD: A prospective study of 21 consecutive eyes (21 patients) was performed. Sufficient antibacterial, antifungal, or antiviral agents were applied to eradicate causative organisms before permanent or temporary amniotic membrane transplantation, or a combination of the two in few patients. The amniotic membrane was soaked in antiinfective agents before transplantation in all cases. RESULTS: After amniotic membrane transplantation, follow-up times ranged from 4 to 28 months (mean, 18 months). Clinical indications included Staphylococcus species (four cases), Pseudomonas species (five cases), Acanthamoeba species (three cases), fungus (two cases), and herpesvirus (seven cases). The corneal surface was healed successfully and recurrences of microbial infection were not noted in any case. Visual acuity was improved in cases that were nonscarring or after additional penetrating keratoplasty. CONCLUSION: Amniotic membrane transplantation seems to be a useful adjunctive surgical procedure for the management of infectious corneal ulcer by promoting wound healing and reducing inflammation.
Asunto(s)
Queratitis por Acanthamoeba/cirugía , Amnios/trasplante , Úlcera de la Córnea/cirugía , Infecciones Bacterianas del Ojo/cirugía , Queratitis Herpética/cirugía , Queratitis por Acanthamoeba/parasitología , Adolescente , Adulto , Anciano , Úlcera de la Córnea/microbiología , Úlcera de la Córnea/parasitología , Úlcera de la Córnea/virología , Infecciones Bacterianas del Ojo/microbiología , Femenino , Humanos , Queratitis Herpética/microbiología , Masculino , Persona de Mediana Edad , Procedimientos Quirúrgicos Oftalmológicos , Estudios Prospectivos , Agudeza Visual , Cicatrización de HeridasRESUMEN
PURPOSE: Stromal herpes simplex virus keratitis (HSK) is an immune-mediated disease. Previous studies have indicated that T cells, neutrophils, and macrophages contribute to the tissue damage in HSK. It has been shown that human amniotic membrane promotes epithelial wound healing and has diverse anti-inflammatory effects. In this study, the effect of amniotic membrane transplantation (AMT) on corneal wound healing and on inflammation in mice with necrotizing HSK was examined. METHODS: BALB/c mice were corneally infected with 10(5) plaque-forming units (PFU) of HSV-1 (KOS strain). In 16 mice that exhibited severe ulcerating HSK, the cornea was covered with a preserved human amniotic membrane as a patch. Corneas in 16 infected mice remained uncovered and served as a control. On days 2 and 7 after surgery, the amniotic membrane was removed (eight mice in each group), the HSV-1-infected cornea was evaluated clinically, and the eye was enucleated. Tissue sections were analyzed histologically for epithelialization and cellular infiltration and immunohistochemically with anti-CD3 mAb to T cells, anti-CD11b mAb to both macrophages and neutrophils, or anti-F4/80 mAb to macrophages. RESULTS: Profound regression of corneal inflammation and rapid closure of epithelial defects were observed clinically within 2 days in the amniotic membrane-covered eyes, whereas HSV-1 keratitis and ulceration progressed in all mice in the control group (P < 0.001). Histologically, corneal edema and inflammatory infiltration, and immunohistochemically the number of CD3(+), CD11b(+), and F4/80(+) cells in the cornea were markedly decreased at 2 and 7 days after amniotic membrane application, compared with the uncovered control corneas (P < 0.001). CONCLUSIONS: AMT promotes rapid epithelialization and reduces stromal inflammation and ulceration in HSV-1 keratitis. AMT in mice with HSV necrotizing stromal keratitis appears to be a useful model for investigating the effect and the action mechanism of human amniotic membrane.
Asunto(s)
Amnios/trasplante , Córnea/cirugía , Herpesvirus Humano 1/fisiología , Queratitis Herpética/cirugía , Cicatrización de Heridas , Animales , Complejo CD3/análisis , Córnea/inmunología , Córnea/microbiología , Humanos , Técnicas para Inmunoenzimas , Queratitis Herpética/inmunología , Queratitis Herpética/microbiología , Antígeno de Macrófago-1/análisis , Macrófagos/inmunología , Ratones , Ratones Endogámicos BALB C , Neutrófilos/inmunología , Linfocitos T/inmunologíaRESUMEN
Fungi belonging to the genus Acremonium Link ex Fries 1821 are ubiquitous environmental contaminants and soil saprophytes, but are infrequent pathogens in humans. These filamentous fungi (previously known as Cephalosporium) are an uncommon cause of mycotic keratitis. As in the case of other filamentous fungi, corneal trauma with contaminated matter is the most frequent risk factor for the infection. We report in this paper a case of keratomycosis caused by Acremoniumpotronii, in a patient with a history of herpetic keratitis. Medical treatment with amphotericin B was unsuccessful and the infection eventually resolved with penetrating keratoplasty.
Asunto(s)
Acremonium/aislamiento & purificación , Infecciones Fúngicas del Ojo/complicaciones , Herpesvirus Humano 1/aislamiento & purificación , Queratitis Herpética/complicaciones , Queratitis Herpética/microbiología , Micosis/complicaciones , Adulto , Anticuerpos Antivirales/análisis , Antifúngicos/uso terapéutico , Sustancia Propia/microbiología , Diagnóstico Diferencial , Infecciones Fúngicas del Ojo/microbiología , Infecciones Fúngicas del Ojo/terapia , Herpesvirus Humano 1/inmunología , Humanos , Queratitis Herpética/terapia , Queratoplastia Penetrante , Masculino , Micosis/microbiología , Micosis/terapiaAsunto(s)
Trasplante de Córnea/efectos adversos , Herpesvirus Humano 1/crecimiento & desarrollo , Queratitis Herpética/etiología , Terapia por Láser/efectos adversos , Activación Viral , 2-Aminopurina/administración & dosificación , 2-Aminopurina/análogos & derivados , 2-Aminopurina/uso terapéutico , Antivirales/administración & dosificación , Antivirales/uso terapéutico , Córnea/microbiología , Córnea/cirugía , Famciclovir , Femenino , Estudios de Seguimiento , Humanos , Hiperopía/cirugía , Queratitis Herpética/tratamiento farmacológico , Queratitis Herpética/microbiología , Queratotomía Radial , Persona de Mediana Edad , Miopía/cirugía , Trifluridina/administración & dosificación , Trifluridina/uso terapéutico , Agudeza VisualRESUMEN
The promoters of the latency-associated transcripts (LATs) of herpes simplex virus type 1 (HSV-1) strains KOS and McKrae were compared to examine their influence upon the reactivation phenotypes of these two strains. Unlike strain KOS, McKrae is readily reactivable using in vivo reactivation models. We found greater than 96% sequence conservation between KOS and McKrae in the LATs promoter region, and both promoters showed equivalent basal and inducible activities. An inter-strain recombinant (termed MK13) was constructed in which the LATs promoter of HSV-1 McKrae was recombined into the background of HSV-1 strain KOS. In a murine u.v. light-induced reactivation model, virus shedding was detected by eye swabbing in two of 44 (5%) mice infected with KOS, 20 of 42 (48%) mice infected with McKrae and none of 45 (0%) mice infected with MK13. These data show that the LATs promoters of these viruses are structurally and functionally similar and that transfer of the LATs promoter from McKrae into KOS is insufficient to confer a reactivatable phenotype.
Asunto(s)
Herpesvirus Humano 1/genética , Queratitis Herpética/microbiología , Regiones Promotoras Genéticas , Transcripción Genética , Replicación Viral , Animales , Secuencia de Bases , Bucladesina/farmacología , Cloranfenicol O-Acetiltransferasa/biosíntesis , Colforsina/farmacología , Secuencia Conservada , Biblioteca Genómica , Herpesvirus Humano 1/patogenicidad , Herpesvirus Humano 1/fisiología , Ratones , Datos de Secuencia Molecular , Células PC12 , Recombinación Genética , Mapeo Restrictivo , Homología de Secuencia de Ácido Nucleico , TransfecciónRESUMEN
PURPOSE: To assess the suitability of the rat for studies of ocular infection with herpes simplex virus (HSV). METHODS: LEW, AO, DA, PVG, and (DAxLEW)F1 x LEW backcross generation rats, 7 to 9 weeks of age, were inoculated with HSV-1 McKrae. The course of primary disease was assessed by clinical observation using a slit lamp. Infectious virus was assayed in ocular and nervous tissue, and the incidence of latent infection was determined. RESULTS: LEW and AO strains were the most susceptible. All LEW rats died after an inoculum of 4 x 10(2) plaque-forming units (pfu) and developed severe corneal disease and uveitis. In contrast, all PVG rats survived 10(4) pfu, 60% survived 4 x 10(4) pfu, and eye disease was restricted to epithelial lesions, sometimes accompanied by mild stromal haze. This resolved, even in animals that developed central nervous system disease. The DA strain showed intermediate susceptibility. Resistance was dominant because disease in backcross generation (DA x LEW)F1 x LEW rats resembled that of the DA rather than the LEW strain. Resistance appeared to be linked to coat color (P < 0.001) rather than to major histocompatibility complex (MHC) type. Chronic stromal disease did not occur in survivors (DA, PVG, and hybrid strains only). CONCLUSIONS: The susceptibility of rat strains to infection of the cornea with HSV varies, and, as with mice, resistance seems to be controlled by non-MHC genes. Rats may prove useful for immunologic studies. Virus reactivation will be the subject of a future report.
Asunto(s)
Herpesvirus Humano 1/fisiología , Queratitis Herpética/microbiología , Animales , Encefalopatías/genética , Encefalopatías/microbiología , Córnea/inmunología , Córnea/microbiología , Susceptibilidad a Enfermedades , Femenino , Inmunidad Innata , Queratitis Herpética/genética , Queratitis Herpética/inmunología , Ratas , Ratas Endogámicas , Latencia del Virus , Replicación ViralRESUMEN
The role of nerve growth factor (NGF) in the modulation of herpes simplex virus (HSV) latency and reactivation was investigated in a mouse model. To determine whether NGF depletion would reactivate latent virus, concentrated anti-NGF serum antibodies were administered intraperitoneally to latently infected mice for 9 consecutive days. To determine whether NGF given prophylactically could suppress UV-B-induced viral reactivation, mice were irradiated with UV-B while being treated with NGF using diverse regimes over a 4-day period. Following intraperitoneal administration of anti-NGF antibodies, viral shedding was detected in a small number (10%) of mice, but it was not possible to pharmacologically suppress UV-B-induced viral reactivation with NGF. It would appear, therefore, that HSV latency in neurons innervating the cornea can be sustained and disrupted by physiological factors independent of NGF levels.
Asunto(s)
Herpesvirus Humano 1/crecimiento & desarrollo , Queratitis Herpética/microbiología , Factores de Crecimiento Nervioso/fisiología , Activación Viral/efectos de los fármacos , Animales , Femenino , Herpesvirus Humano 1/efectos de la radiación , Queratitis Herpética/patología , Ratones , Ratones Endogámicos , Recurrencia , Ganglio Cervical Superior/microbiología , Ganglio Cervical Superior/patología , Ganglio del Trigémino/microbiología , Ganglio del Trigémino/patología , Rayos Ultravioleta , Activación Viral/efectos de la radiación , Latencia del Virus/fisiología , Esparcimiento de Virus/fisiologíaRESUMEN
We have used a mouse model system and the corneal route of inoculation to examine the issue of extra-neuronal persistence of herpes simplex virus type 1 (HSV-1). HSV-1 strain F DNA and inflammatory lesions were detected in corneal tissue of mice at 5, 11, 23, 37 and 60 days post-infection (p.i.) Viral DNA was localized by in situ PCR to epithelial cells and less frequently to cells in the stroma of the cornea. Viral proteins were not detected in the cornea and virus could not be isolated from tissue homogenates after 11 days p.i. even though histopathological lesions became progressively more severe at 37 and 60 days p.i. The DNA-containing cells were usually adjacent to the sites of inflammation or within these sites in the chronic stage (23, 37 and 60 days p.i.). In contrast to strain F, persistence of HSV-1 strain KOS DNA and inflammatory lesions were not detected after 11 days p.i.; this result suggests that the long-term persistence of HSV-1 DNA and the development of inflammatory lesions are virus strain-dependent. We tested for the possibility of transgenic HSV-1 immediate early gene (ICP4) promoter activity in chronically infected corneas of transgenic mice containing the ICP4 promoter fused to the bacterial beta-galactosidase coding sequence. Our results indicated that the chimeric transgene was expressed in the cornea at 5, 11, 23, 37 and 41 days p.i. Possible explanations for these results and mechanisms for the generation of the chronic inflammatory lesions are discussed. The properties of chronic HSV infections in the cornea may be similar to those which have been described for persistent or defective viral infections in other systems.
Asunto(s)
Enfermedades de la Córnea/microbiología , Herpesvirus Humano 1/patogenicidad , Queratitis Herpética/microbiología , Animales , Antígenos Virales/análisis , ADN Viral/análisis , Epitelio/microbiología , Ojo/microbiología , Regulación Viral de la Expresión Génica , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C3H , Ratones Endogámicos C57BL , Ratones Transgénicos , Neuronas/microbiología , Regiones Promotoras Genéticas , ARN Mensajero/genéticaRESUMEN
Unilateral intravitreal inoculation of herpes simplex virus type 1 (HSV) induced bilateral retinitis and encephalitis in rats. In a previous paper, we demonstrated immunohistochemical localization of HSV in the ipsilateral retina. In this paper, we report immunohistochemical localization of antigen to HSV in the brain and opposite eye. HSV was found in the optic chiasm, and bilaterally in the suprachiasmatic nuclei, lateral geniculate nuclei, superior colliculus, pretectum, and visual cortex. HSV was also found in the retinal ganglion cells, neural cells in the inner nuclear layer, and Müller cells in the opposite eye. We demonstrated HSV infection in the retina of the opposite eye via the optic nerve, visual pathway, and nuclei, but not through the blood stream.
Asunto(s)
Encéfalo/microbiología , Herpesvirus Humano 1/aislamiento & purificación , Queratitis Herpética/microbiología , Nervio Óptico/microbiología , Retina/microbiología , Retinitis/microbiología , Animales , Antígenos Virales/análisis , Inmunohistoquímica , Masculino , Ratas , Ratas Sprague-DawleyRESUMEN
We evaluated the epithelializing-promoting effect of a concentration of 0.005 mg/ml murine epidermal growth factor (mEGF), topically administered four times daily, on the herpes simplex virus 1 (HSV-1) corneal ulcers of the rabbit eye. The severity of the herpetic lesions was evaluated clinically, after the time course of the severity of epithelial keratitis, conjunctivitis, iritis, and stromal disease, for 14 days. A histological assessment was performed in the middle and at the end of the follow-up. The stromal keratitis of the mEGF-treated group was significantly more severe than the keratitis exhibited by the placebo-treated rabbits (Y = X3 X2*X4; p = 0.0001). There were no significant differences in the degree of conjunctivitis, epithelial keratitis, iritis, and virus shedding between these groups. No evidence of a toxic effect of mEGF or placebo was found in the mock infected rabbit eyes. More studies, using different herpes virus strains and a broad range of murine and human EGF concentrations, are mandatory to ascertain the general significance of these results. Meanwhile, caution is recommended when using mEGF in the presence of an occult or manifest herpetic eye disease.
Asunto(s)
Sustancia Propia/microbiología , Factor de Crecimiento Epidérmico/farmacología , Queratitis Herpética/microbiología , Administración Tópica , Animales , Sustancia Propia/patología , Modelos Animales de Enfermedad , Factor de Crecimiento Epidérmico/efectos adversos , Herpesvirus Humano 1/fisiología , Ratones , Conejos , Distribución Aleatoria , Células Vero , Esparcimiento de VirusRESUMEN
PURPOSE: Fluorescein and rose bengal are dyes used routinely in the examination of the ocular surface. As part of an ongoing search for a superior ophthalmic dye with optimal specificity and sensitivity and a lack of interference with subsequent viral cultures, and as part of studies that use chemical dyes to understand better the pathophysiology of ocular surface disorders, the staining characteristics and antiviral activity of sulforhodamine B and lissamine green B were investigated. METHODS: Staining of rabbit corneal epithelial cell cultures by sulforhodamine B and lissamine green B was compared to that of fluorescein and rose bengal. Diffusion of each dye through a collagen gel was measured. Uptake of lissamine green B by herpes simplex virus type 1 (HSV-1)-infected Vero cell cultures was compared at several times postinfection. The effect of sulforhodamine B and lissamine green B on HSV-1 plaque formation in Vero cells was determined. The cellular toxicity of sulforhodamine B and lissamine green B in vitro was examined by a quantitative 14C-amino acid uptake assay and by a qualitative cell viability assay. Finally, the effect of sulforhodamine B and lissamine green B on viral replication was compared in vivo with that of rose bengal in a rabbit model of herpetic epithelial keratitis. RESULTS: Rose bengal vividly stained cell monolayers of explant cultures of rabbit corneal epithelium. By light microscopy, sulforhodamine B and lissamine green B, like fluorescein, did not stain the epithelial cells, but did stain the corneal explant stroma. Pretreatment of epithelial cells with 0.25% trypsin for 5 minutes failed to induce dye uptake; however, pretreatment with 0.5% Triton X-100 for 5 minutes resulted in nuclear staining by lissamine green B, but not sulforhodamine B. When added to a collagen gel, the relative diffusion rate was fluorescein > lissamine green B > sulforhodamine B > rose bengal. By spectrophotometric analysis, HSV-1-infected and uninfected Vero cells bound equivalent amounts of lissamine green B until late in infection, when infected cells took up more dye (P < 0.001). A direct neutralization assay showed that 0.06% lissamine green B or 0.5% sulforhodamine B reduced HSV-1 plaque formation in Vero cells by greater than 50%, when present at the time of viral adsorption. By a quantitative 14C-amino acid uptake assay, lissamine green B was toxic to Vero cells in a dose-dependent manner, whereas sulforhodamine B was relatively nontoxic at the concentrations tested. By a cell viability assay, however, neither dye showed significant cellular toxicity. In a rabbit model of herpetic epithelial keratitis, rose bengal significantly reduced viral replication and recovery, whereas sulforhodamine B and lissamine green B had no effect. CONCLUSIONS: Neither sulforhodamine B nor lissamine green B stain healthy, normal cells. Lissamine green B stains membrane-damaged epithelial cells, but sulforhodamine B does not. Both sulforhodamine B and lissamine green B stain corneal stroma. Lissamine green B inhibits HSV-1 plaque formation at low concentrations of dye in vitro, which correlates with suppression of cellular metabolism as demonstrated by a 14C-amino acid uptake assay, but does not affect cell viability. Neither sulforhodamine B nor lissamine green B inhibit viral replication or recovery in vivo.
Asunto(s)
Herpesvirus Humano 1/efectos de los fármacos , Colorantes Verde de Lisamina/farmacología , Rodaminas/farmacología , Coloración y Etiquetado , Animales , Córnea/efectos de los fármacos , Córnea/microbiología , Córnea/patología , Modelos Animales de Enfermedad , Herpesvirus Humano 1/fisiología , Queratitis Herpética/microbiología , Queratitis Herpética/patología , Colorantes Verde de Lisamina/farmacocinética , Colorantes Verde de Lisamina/toxicidad , Conejos , Rodaminas/farmacocinética , Rodaminas/toxicidad , Coloración y Etiquetado/métodos , Células Vero , Replicación Viral/efectos de los fármacosRESUMEN
We investigated the use of the polymerase chain reaction for detecting genomes of herpes simplex virus, varicella-zoster virus, and cytomegalovirus from tear film of patients with clinically diagnosed herpes simplex virus keratitis. Using the polymerase chain reaction with a herpes simplex virus detection sensitivity adjusted to 1.0 plaque-forming units/ml, we detected herpes simplex virus genomic sequences in 12 of 12 epithelial keratitis specimens, two of six stromal keratitis specimens, but in none of 20 normal specimens. Neither varicella-zoster virus nor cytomegalovirus genomic sequences were detected in any sample. These results suggest that polymerase chain reaction quickly performed with reduced sensitivity is useful as a diagnostic tool for confirming clinical observations.
Asunto(s)
ADN Viral/análisis , Simplexvirus/genética , Lágrimas/microbiología , Secuencia de Bases , Citomegalovirus/genética , Herpesvirus Humano 3/genética , Humanos , Queratitis Herpética/microbiología , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa , Sensibilidad y Especificidad , Simplexvirus/aislamiento & purificaciónRESUMEN
Rose bengal and fluorescein are photosensitive dyes in widespread use in the evaluation of ocular surface diseases, including herpes simplex virus (HSV) keratitis. These dyes have recently been shown to penetrate living cells, and rose bengal was previously reported to possess antiviral activity. Several experiments reported herein suggest that these dyes do possess the potential for potent antiviral activity against extracellular virus, but only in the presence of light. Rose bengal is substantially more effective in vitro than fluorescein, and the effect is greater with increasing concentration of dye and duration of light exposure. Electron microscopic evaluation of treated virus showed no structural difference from untreated virus, in spite of 4- to 5-log decreases in virus titer. Intracellular virus was found to be markedly resistant to photoinactivation. In a rabbit model of acute primary HSV keratitis, daily application of topical rose bengal followed by light exposure had no therapeutic effect, although an adverse effect on culture sensitivity testing was seen.