In vitro amoebicidal effects of arabinogalactan-based ophthalmic solution.

The main corneal infections reported worldwide are caused by bacteria and viruses but, recently, the number of Acanthamoeba keratitis (AK) cases has increased. Acanthamoeba genus is an opportunistic free living protozoa widely distributed in environmental and clinical sources, with two life-cycle stages: the trophozoite and the cyst. AK presents as primary symptoms eye redness, epithelial defects, photophobia and intense pain. An early diagnosis and an effective treatment are crucial to avoid blindness or eye removal but, so far, there is no established treatment to this corneal infection. Diverse research studies have reported the efficacy of commercialized eye drops and ophthalmic solutions against the two life cycle stages of Acanthamoeba strains, that usually present preservatives such as Propylene Glycol of Benzalkonium chloride (BAK). These compounds present toxic effects in corneal cells, favouring the inflammatory response in the so sensitive eye tissue. In the present work we have evaluated the efficacy of nine proprietary ophthalmic solutions with and without preservatives (ASDA Dry Eyes Eyedrops, Miren®, ODM5®, Ectodol®, Systane® Complete, Ocudox®, Matrix Ocular®, Alins® and Coqun®) against the two life cycle stages of three Acanthamoeba strains. Our work has demonstrated the high anti-Acanthamoeba activity of Matrix Ocular®, which induces the programmed cell death mechanisms in Acanthamoeba spp. trophozoites. The high efficacy and the absence of ocular toxic effects of Matrix Ocular®, evidences the use of the Arabinogalactan derivatives as a new source of anti-AK compounds.

Queratoconjuntivitis por Vittaforma corneae: una enfermedad emergente entre los viajeros que regresan del sudeste asiático / Vittaforma Corneae keratoconjunctivitis: An emerging pathology among travelers returning from Southeast Asia

La queratoconjuntivitis por Microsporidium es un cuadro infrecuente. Se ha asociado a brotes epidémicos en Asia relacionados con la exposición a agua o tierra contaminada. Habitualmente estos cuadros son autolimitados y leves, pero pueden tener evoluciones prolongadas. Presentamos el caso de una paciente de 29 años que comenzó con dolor, enrojecimiento, visión borrosa en su ojo derecho tras su vuelta de un viaje a Singapur y que no mejoró tras un tratamiento convencional frente a conjuntivitis. Fue diagnosticada de queratoconjuntivitis por Microsporidium mediante PCR y tinción con PAS del epitelio corneal. El tratamiento inicial fue desbridamiento epitelial, albendazol oral y voriconazol, levofloxacino y propamidina, pero la enfermedad conjuntival y corneal no cedió hasta que si introdujeron corticoides tópicos 5 meses después para tratar la limbitis. Debemos tener la sospecha de queratitis por Microsporidium en casos de queratitis unilateral y conjuntivitis, sobre todo en pacientes que vuelvan de Asia

Microsporidium keratoconjunctivitis is an very rare disease. It is related to outbreaks in Asia due to exposure to contaminated water or soil. Microsporidium keratoconjunctivitis is a a self-limited disease, but it could have long term courses. We present the case of a 29 year old woman who started with pain, redness and blurred vision after a holiday in Singapore and did not respond to conjunctivitis treatment. PCR sequencing and PAS staining of corneal epithelial biopsy identified Vittaforma corneae as the causative organism. Treatment was initiated with corneal debridement, oral albendazol, and intensive topical voriconazole, levofloxacin and propamidine, but the conjunctival and corneal disease was only resolved 5 months later with the introduction of topical steroids to treat her severe limbitis. Suspicion of Microsporidium keratoconjunctivitis should be raised amongst ophthalmologists in unilateral keratitis with mild conjunctivitis in travelers from Asia

Vittaforma Corneae keratoconjunctivitis: An emerging pathology among travelers returning from Southeast Asia. / Queratoconjuntivitis por Vittaforma corneae: una enfermedad emergente entre los viajeros que regresan del sudeste asiático.

Microsporidium keratoconjunctivitis is an very rare disease. It is related to outbreaks in Asia due to exposure to contaminated water or soil. Microsporidium keratoconjunctivitis is a a self-limited disease, but it could have long term courses. We present the case of a 29 year old woman who started with pain, redness and blurred vision after a holiday in Singapore and did not respond to conjunctivitis treatment. PCR sequencing and PAS staining of corneal epithelial biopsy identified Vittaforma corneae as the causative organism. Treatment was initiated with corneal debridement, oral albendazol, and intensive topical voriconazole, levofloxacin and propamidine, but the conjunctival and corneal disease was only resolved 5 months later with the introduction of topical steroids to treat her severe limbitis. Suspicion of Microsporidium keratoconjunctivitis should be raised amongst ophthalmologists in unilateral keratitis with mild conjunctivitis in travelers from Asia.

Desprendimiento del epitelio pigmentario de la retina en la retinopatía por malaria / Detachment of retinal pigment epithelium in retinopathy due to malaria

CASO CLÍNICO: Varón de 45 años de edad, acude diagnosticado de paludismo con afectación neurológica. Dos meses más tarde refiere metamorfopsia en el ojo izquierdo. En el fondo de ojo se aprecia retinopatía palúdica bilateral, observándose en la tomografía de coherencia óptica (OCT) macular un desprendimiento del epitelio pigmentario (DEP) parafoveal en el ojo izquierdo. Se inicia tratamiento específico antipalúdico, evidenciándose desaparición de la retinopatía. DISCUSIÓN: El Plasmodium falciparum es responsable de la retinopatía en el paludismo neurológico. Debiéndose realizar en estos pacientes una exploración fundoscópica y OCT macular porque está asociado a una mortalidad mayor cuando existe afectación retiniana

CASE REPORT: A 45-year-old man was diagnosed with malaria with neurological involvement. Two months later he referred metamorphopsia in the left eye. Malarial retinopathy was observed in the fundus examination. The Optic Coherence Tomography (OCT) of the macula showed parafoveal pigment epithelium detachment (DEP). Specific anti-malarial treatment was initiated, with the disappearance of the retinopathy being observed. DISCUSSION: Plasmodium falciparum is responsible for the retinopathy in neurological malaria. A funduscopic examination and macular OCT should be performed in these patients, as it is associated with a higher mortality when there is a retinal involvement

Detachment of retinal pigment epithelium in retinopathy due to malaria. / Desprendimiento del epitelio pigmentario de la retina en la retinopatía por malaria.

CASE REPORT: A 45-year-old man was diagnosed with malaria with neurological involvement. Two months later he referred metamorphopsia in the left eye. Malarial retinopathy was observed in the fundus examination. The Optic Coherence Tomography (OCT) of the macula showed parafoveal pigment epithelium detachment (DEP). Specific anti-malarial treatment was initiated, with the disappearance of the retinopathy being observed. DISCUSSION: Plasmodium falciparum is responsible for the retinopathy in neurological malaria. A funduscopic examination and macular OCT should be performed in these patients, as it is associated with a higher mortality when there is a retinal involvement.

Retinopatía MEK. Casos clínicos / MEK Retinopathy. Clinical case reports

CASOS CLÍNICOS: Se presentan 3 casos clínicos de retinopatía MEK asociados al uso de la combinación de cobimetinib y vemurafenib, caracterizados por la alteración del epitelio pigmentario de la retina y desprendimiento neurosensorial. Dos de ellos conservaron la visión de la unidad, el tercero desarrolló un gran desprendimiento neurosensorial bilateral con una agudeza visual final de 0,6 en el ojo derecho y de 0,1 en el izquierdo. DISCUSIÓN: Las nuevas estrategias terapéuticas frente al melanoma cutáneo metastásico condicionan la aparición de alteraciones del epitelio pigmentario de la retina con desprendimientos serosos, lo que obliga a una vigilancia estrecha bajo tomografía de coherencia óptica macular

CASE REPORTS: Three clinical cases are presented of MEK retinopathy associated with the combination of cobimetinib and vemurafenib characterised by alteration of the retinal pigment epithelium and neurosensory detachment. Two of the cases conserved the vision of the unit, and the third developed a large bilateral neurosensory detachment with final visual acuity of 0.6 for the right eye and 0.1 for the left one. DISCUSSION: The new therapeutic strategies against metastatic cutaneous melanoma condition the appearance of alterations of the pigmentary epithelium of the retina with serous detachments, leading to close monitoring with macular optical coherence tomography

MEK Retinopathy. Clinical case reports. / Retinopatía MEK. Casos clínicos.

CASE REPORTS: Three clinical cases are presented of MEK retinopathy associated with the combination of cobimetinib and vemurafenib characterised by alteration of the retinal pigment epithelium and neurosensory detachment. Two of the cases conserved the vision of the unit, and the third developed a large bilateral neurosensory detachment with final visual acuity of 0.6 for the right eye and 0.1 for the left one. DISCUSSION: The new therapeutic strategies against metastatic cutaneous melanoma condition the appearance of alterations of the pigmentary epithelium of the retina with serous detachments, leading to close monitoring with macular optical coherence tomography.

Mácula en cúpula asociada a la retinocoroidosis miópica. Utilidad de la espironolactona / Dome shaped macula associated with myopic retinochoroidosis. Use of spironolactone

Casos clínicos: Se presentan los casos de 3 mujeres de 22, 36 y 55 años de edad con retinocoroidosis miópica bilateral. Las pacientes presentan disminución de agudeza visual unilateral, tonometría y biomicroscopia bilateral normal. En la funduscopia se evidencia maculopatía unilateral, y en la tomografía de coherencia óptica (OCT), mácula en cúpula con desprendimiento neurosensorial (DNS). Iniciado tratamiento con espironolactona, en todos los casos se comprueba mejoría por OCT. Discusión: Se discute el mecanismo etiopatogénico de la mácula en cúpula. La OCT se demuestra como técnica fundamental en el seguimiento de esta patología. Tras la evidencia mostrada, se postula tratamiento inicial con espironolactona (AU)

Cases reports: The cases are presented of three women of 22, 36 and 55 years old with bilateral myopic retinochoroidosis. They had unilateral decreased visual acuity, normal bilateral tonometry, and biomicroscopy. Funduscopy showed bilateral and unilateral myopic maculopathy, and Optical Coherence Tomography (OCT) showed a dome shaped macula with neurosensory detachment. Treatment was started with spironolactone and an improvement by OCT was shown in all cases. Discussion: The etiopathogenic mechanism of the dome shaped macula is discussed. OCT demonstrated to be the fundamental test in the follow-up of this condition. After the evidence shown, initial treatment with spironolactone is suggested (AU)

Dome shaped macula associated with myopic retinochoroidosis. Use of spironolactone. / Mácula en cúpula asociada a la retinocoroidosis miópica. Utilidad de la espironolactona.

CASES REPORTS: The cases are presented of three women of 22, 36 and 55 years old with bilateral myopic retinochoroidosis. They had unilateral decreased visual acuity, normal bilateral tonometry, and biomicroscopy. Funduscopy showed bilateral and unilateral myopic maculopathy, and Optical Coherence Tomography (OCT) showed a dome shaped macula with neurosensory detachment. Treatment was started with spironolactone and an improvement by OCT was shown in all cases. DISCUSSION: The etiopathogenic mechanism of the dome shaped macula is discussed. OCT demonstrated to be the fundamental test in the follow-up of this condition. After the evidence shown, initial treatment with spironolactone is suggested.

The amoebicidal activity of Ziziphus vulgaris extract and its fractions on pathogenic Acanthamoeba trophozoites and cysts.

Acanthamoeba genus includes pathogenic species which are causal agents of a severe sight-threatening infection of the eye known as Acanthamoeba keratitis (AK). Furthermore, the number of AK is worryingly increasing worldwide, mostly in contact lens users. Until present, there is a general failure to reach a fully effective treatment against AK which is mainly due to the amoebic double-walled cyst stage which forms a protective barrier against drugs. Therefore, drug discovery research towards AK treatment is a must. In this study, Ziziphus vulgaris, a native plant of Asian countries, was checked for its activity against Acanthamoeba. For this purpose and in order to determine the in vitro amoebicidal effects of Ziziphus vulgaris aqueous extract and its fractions (chloroformic, remaining aqueous and primary alcoholic) against Acanthmoeba trophozoites and cysts, activity and sensitivity assays were performed. Moreover, the toxic effect of the extract and its fractions was also tested on murine peritoneal macrophages using a colorimetric tetrazolium salt (MTT) test. The obtained results showed that the chloroformic fraction presented a higher anti-Acanthamoeba activity when compared to the other fractions (Trophozoites/cysts were eliminated, when incubated in a concentration of 50 mg/ml of the fraction, after 24 hours). The calculated active concentrations against Acanthamoeba of these extracts did not shown any high cytotoxicity levels.This study suggests that the Ziziphus vulgaris chloroformic fraction, may present compounds with relevance for the development of novel antiAcanthamoeba drugs.

Isolation of Naegleria fowleri from a domestic water tank associated with a fatal encephalitis in a 4 month-old Venezuelan child.

This study describes the association of household water system contamination with the pathogenic Free-Living Amoeba (FLA) Naegleria fowleri and a case of fatal Primary Amoebic Meningoencephalitis (PAM) in a child from the state of Monagas in Venezuela. Amoebae were initially identified by microscopy from a sample of cerebrospinal fluid (CSF) from the child. Direct DNA extraction and specific PCR/sequencing for N. fowleri was also carried out from the same CSF sample. In order to determine a possible environmental source of infection, water samples from the water tank of the child's home and also water bodies recently visited by the child and his family, were examined for the presence of N. fowleri by culture and PCR/sequencing. The results obtained from the collected water samples revealed that only the water tank of the house was positive for N. fowleri. PCR/sequencing showed that the strains isolated from the patient and the water tanks were 100 % identical. Therefore, the house water tank was confirmed as the source of infection in this case, possibly as a result of the occasional immersion of the child´s head under the water while bathing. This case highlights a novel source of thermally polluted water and another threat of N. fowleri infection.

An epidemiological study of A. cantonensis in Jamaica subsequent to an outbreak of human cases of eosinophilic meningitis in 2000.

The infection status of angiostrongylosis in Jamaica was assessed in wild rats and molluscs in the 5 years following the major outbreak of eosinophilic meningitis (EM) in 2000. Parasitological analyses of 297 Rattus rattus and 140 Rattus norvegicus, and 777 terrestrial molluscs from all 14 Parishes on the island revealed Angiostrongylus cantonensis in 32·0% of the rats and in 12·5% of the molluscs. Multivariate analyses confirmed that A. cantonensis occurred significantly more frequently in R. rattus (Odds Ratio [OR] = 1·76), while mean infection intensity in R. rattus was also significantly higher (16·8) than R. norvegicus (11·3) (Mann-Whitney U-test: P = 0·01). Third-stage larvae of A. cantonensis were detected in 29% of 86 Pleurodonte spp.; in 20% of five Poteria spp.; in 18·7% of 369 Thelidomus asper; in 11% of 18 Sagda spp.; and in 6% of 24 veronicellid slugs. Most rodent infections occurred in Northeastern Jamaica (OR = 11·66), a region where infected molluscs were also abundant. Given the prevalence of A. cantonensis infection in rats has significantly increased since the 2000 outbreak, and that a survey of human infections revealed at least ten autochthonous cases in the last 15 years, angiostrongylosis persists as an important zoonosis in Jamaica.

Presence of Balamuthia mandrillaris in hot springs from Mazandaran province, northern Iran.

Balamuthia mandrillaris is an opportunistic free-living amoeba that has been reported to cause cutaneous lesions and Balamuthia amoebic encephalitis. The biology and environmental distribution of B. mandrillaris is still poorly understood and isolation of this pathogen from the environment is a rare event. Previous studies have reported that the presence of B. mandrillaris in the environment in Iran may be common. However, no clinical cases have been reported so far in this country. In the present study, a survey was conducted in order to evaluate the presence of B. mandrillaris in hot-spring samples of northern Iran. A total of 66 water samples were analysed using morphological and molecular tools. Positive samples by microscopy were confirmed by performing PCR amplification of the 16S rRNA gene of B. mandrillaris. Sequencing of the positive amplicons was also performed to confirm morphological data. Two of the 66 collected water samples were positive for B. mandrillaris after morphological and molecular identification. Interestingly, both positive hot springs had low pH values and temperatures ranging from 32 °C to 42 °C. Many locals and tourists use both hot springs due to their medicinal properties and thus contact with water bodies containing the organism increases the likelihood of infection. To the best of our knowledge, this is the first report on the isolation of B. mandrillaris from hot-spring sources related to human activity. Therefore, B. mandrillaris should be considered as a possible causative agent if cases of encephalitis are suspected following immersion in hot springs in addition to Acanthamoeba and Naegleria.

Gastrointestinal helminths of wild hogs and their potential livestock and public health significance in Jamaica.

An investigation into the potential for transmission of gastrointestinal helminths from wild hogs to livestock and humans was prompted by concerns of recreational wild-hog hunting in the Caribbean region and the recent practice, by livestock farmers in Jamaica, of co-rearing wild and domesticated swine. Thirty-one wild hogs from the Hellshire Hills, a dry limestone forest in southern Jamaica, were necropsied during the period June 2004 to August 2006. Thirteen of the captured animals were male and 18 female. Four species of adult helminths were recovered from the gastrointestinal tracts of the wild hogs: Hyostrongylus rubidus (77%), Globocephalus urosubulatus (48%), Oesophagostomum dentatum (42%) and Macroacanthorhynchus hirudinaceus (77%). Two (6.2%), ten (32.2%) and 18 (58.0%) hogs harboured one, two and three species of helminths, respectively. Mean infection intensities varied from 8.1 for M. hirudinaceus, to 115.5 for O. dentatum. There was no association between any of the recovered helminths and sex of the host; however, a multivariate analysis indicated a positive association between the prevalence of G. urosubulatus and host age (odds ratio (OR) = 6.517). Domesticated hogs co-reared with wild hogs are potentially at risk of infection with all four helminths, while wild-hog hunters and pig farmers may be exposed to M. hirudinaceus.

High occurrence of potentially-pathogenic free-living amoebae in tap water and recreational water sources in South-West Iran.

The present study was conducted to evaluate the presence of potentially pathogenic free living amoebae in drinking and recreational water sources in south-western Iran. From 75 collected water samples, 40 samples (53.3%) were positive for free living amoebae identified using morphological tools. Interestingly, all recreational waters in Ilam city included in the present study were positive for Acanthamoeba, Vahlkampfidae and Vermamoeba. Thirty percent of tap water samples in Ahvaz city were also positive for potentially pathogenic Free Living Amoebae. Moreover, the three genera identified in the present study have been previously reported as keratitis causative agents in Iran. The present research highlights the need to improve filtration methods for tap waters and to establish awareness in recreational water sources in Iran, in order to prevent Free Living Amoebae related infections. To the best of our knowledge, this is the first report of the isolation of potentially pathogenic free living amoebae including Acanthamoebae, Vermamoebae and Vahlkampfiids in the South-West of Iran.

Balamuthia mandrillaris therapeutic mud bath in Jamaica.

Balamuthia mandrillaris is an emerging cause of encephalitis in humans. The transmission dynamics are poorly understood due to the high fatality rate and the sporadic nature of cases. Seventy-two soil samples were collected from beaches and the banks of lagoons, rivers, ponds, mineral springs and streams from across Jamaica and assayed for the presence of B. mandrillaris. Seventy-nine sites were sampled and the mitochondrial 16S rDNA gene of B. mandrillaris was amplified and sequenced to confirm the presence of the amoeba. One isolate of B. mandrillaris was recovered from soil from mineral spring which hosts an informal therapeutic mud bath business. Although B. mandrillaris is less frequently isolated from soil than other free-living amoebae, rubbing mud containing the organism onto the skin increases the likelihood of exposure and infection. This first report on the isolation of B. mandrillaris in the Caribbean and its presence in soil where human contact is likely warrants further investigation using serological methods to elucidate exposure patterns.

Evaluation of the in vitro activity of commercially available moxifloxacin and voriconazole eye-drops against clinical strains of Acanthamoeba.

PURPOSE: Acanthamoeba is an opportunistic pathogen which is the causal agent of a sight-threatening ulceration of the cornea known as "Acanthamoeba keratitis" (AK) and, more rarely, an infection of the central nervous system called "granulomatous amoebic encephalitis" (GAE). The symptoms of AK are non-specific, and so it can be misdiagnosed as a viral, bacterial, or fungal keratitis. Furthermore, current therapeutic measures against AK are arduous, and show limited efficacy against the cyst stage of Acanthamoeba. Moxifloxacin, a fourth generation fluoroquinolone, has been used with other drugs to treat GAE, but its efficacy as a treatment for AK is not known. Voriconazole has been used to treat AK; however, its cysticidal efficacy is not known. Both drugs are commercially available as eye-drops. The aim of this study was to evaluate the in-vitro activity of these eye-drops against Acanthamoeba compared to two reference drugs (chlorhexidine and amphotericin B) which are currently used to treat AK and GAE. METHODS: The sensitivity of two clinical and one type strain of Acanthamoeba to the commercial concentrations of the four drugs was evaluated with a colorimetric assay. Mature cysts were incubated with voriconazole to determine their sensitivity to this drug. The effects on cell proliferation and cell toxicity were determined using standard procedures with commercial kits. RESULTS: The four compounds were active against the Acanthamoeba strains in this study. Although it prevented encystation, moxifloxacin's amoebicidal activity was low. Voriconazole activity was greater than that of the other drugs, even at a concentration lower than in commercial eye drops. It was effective against cysts and decreased cell proliferation, with low cellular cytotoxicity. CONCLUSION: Voriconazole could be used against AK as a first-line treatment or in combination. Moxifloxacin is an interesting adjuvant to consider as it is effectively prevents encystation of the amoeba which often complicates infection resolution. In addition, moxifloxacin is effective in preventing secondary bacterial infections.

Pathogenic Acanthamoeba strains from water sources in Jamaica, West Indies.

In 2004, samples of tap water and of river and sea water associated with human activities were collected in Jamaica, West Indies, and checked for free-living Acanthamoeba. The morphologies of the cysts and trophozoites observed and the results of PCR-based amplifications with a genus-specific primer pair were used to identify the Acanthamoeba isolates. The potential of each isolate as a human pathogen was then evaluated using thermotolerance and osmotolerance assays and two PCR-based assays for Acanthamoeba pathogenesis. Acanthamoeba were identified in 36.1%, 26.4% and 49.6% of the tap-, river- and sea-water samples collected, respectively. Pathogenic potential was shown by 60.0% of the Acanthamoeba strains isolated from tap water, 68.4% of the strains from river water, and 40.4% of the seawater strains. Sequencing of ribosomal DNA revealed the T1, T2, T4, T5, T7, T9 and T11 genotypes. Isolates of the T4 genotype were collected from tap, rain and sea water and, as expected, exhibited the most pathogenic traits; most were osmotolerant, thermotolerant and expressing extracellular serine protease. This is the first study of the occurrence and distribution of Acanthamoeba in water in the West Indies, and the results confirm the presence of potentially pathogenic strains in Jamaica.

A specific primer pair for the diagnosis and identification of Acanthamoeba astronyxis by random amplified polymorphic DNA-polymerase chain reaction.

Random amplified polymorphic DNA (RAPD) is a useful tool for species identification. The obtained band patterns can be used for specific primer pair design that is useful for species identification. In this study, a distinctive 485-bp band in Acanthamoeba astronyxis band patterns was found, using the OPC20 primer (ACTTCGCCAC). The band specificity was confirmed by hybridization, using it as a probe, against all OPC20 amplifications from different Acanthamoeba species. Once the fragment was sequenced, we used it to design a specific primer pair that was useful for the identification of different isolates as A. astronyxis species.