Acanthamoeba spp. en piscinas cubiertas de la ciudad de Bahía Blanca, provincia de Buenos Aires, Argentina / Acanthamoeba spp. in indoor swimming pools in Bahía Blanca city, Buenos Aires province, Argentina

Acanthamoeba ha sido aislada de numerosos hábitats incluyendo piletas de natación. Estudiar su distribución es importante ya que algunas especies causan enfermedad en el hombre. El objetivo del presente trabajo fue la búsqueda, aislamiento y caracterización de protozoos del género Acanthamoeba en piscinas cubiertas de la ciudad de Bahía Blanca, Argentina, en las cuatro estaciones del año durante 2007-2008. Se estudiaron 7 piscinas y en cada una se tomaron cuatro muestras: fondo, superficie, raspado de pared y para análisis bacteriológico. Las muestras se analizaron por observación directa y por cultivo en agar no nutritivo a 37 °C y a 42 °C. La identificación genérica se realizó de acuerdo con las características morfológicas de quistes y trofozoítos y para identificar Naegleria se realizó la prueba de transformación ameboflagelar. En 5 de las 7 piscinas se aislaron amebas de vida libre al menos en una época del año. La prueba de transformación ameboflagelar resultó negativa, descartando al género Naegleria. Todos los aislamientos correspondieron al género Acanthamoeba Grupos II y III de Pussard y Pons. Si bien la eliminación de Acanthamoeba en las aguas de natatorios resulta muy difícil por tratarse de un protozoario ubicuo y sumamente resistente a los desinfectantes comúnmente utilizados, se recomienda una limpieza profunda de las piscinas que minimice los riesgos de infección.

Acanthamoeba spp. has been isolated from many habitats, including swimming pools. Investigations on its distribution are relevant because many of its species cause human diseases. The aim of the present work was to investigate, isolate and characterize protozoan of the genus Acanthamoeba from indoor swimming pools in Bahía Blanca, Argentina, in the four seasons, during the 2007-2008 period. Seven pools were studied and samples were collected from the bottom, surface and wall by scraping them. Besides, samples for bacteriological analysis were taken from each pool. The samples were analyzed by direct observation and by culturing on non nutritive agar at 37 °C and at 42 °C. The generic identification was performed according to the morphologic characteristics of cysts and trofozoites, while the amoebo-flagellate transformation test was carried out to identify Naegleria. Free-living amoebas were isolated from five of the seven swimming pools examinated, in at least one season. Naegleria genus was not found as the amoebo-flagellate transformation tests were negative in all samples. All the isolations corresponded to the genus Acanthamoeba belonging to Pussard & Pons morphological Groups II and III. Although elimination of Acanthamoeba from the water is difficult because it is a ubiquitous protozoan. which is highly resistant to the commonly used disinfectants, a thorough cleaning of the pools to minimize the risks of infection is recommended.

Acanthamoeba spp: ecoepidemiología, biología, ultraestructura, patogénesis y diagnóstico en el hombre / Acanthamoeba: ecoepidemiology, biology, ultrastructure, pathogenesis and diagnosis in humans

En el presente trabajo presentamos nuestros estudios referidos al protozoo Acanthamoeba, desde lo morfológico, diagnóstico de laboratorio, patología en el ser humano, aspectos epidemiológicos y resistencia al cloro. Se aisló Acanthamoeba en el 28.6% de los tanques domiciliarios de agua, en el 71% de piscinas cubiertas, en la totalidad de las muestras del arroyo Napostá, que cruza la ciudad de Bahía Blanca, y no fue detectada en el agua de la red. Con referencia a la acción quisticida del cloro, se logró inhibir el crecimiento recién a las tres horas de exposición en 60 ppm de cloro. Realizamos el primer aislamiento de un paciente con queralitis, en Bahía Blanca. Hasta el momento no aislamos Acanthamoeba de otras muestras biológicas. Con referencia a los aspectos morfológicos, documentamos la ultraestructura de las cepas regionales de Acanthamoeba y realizamos la tipificación molecular de la cepa aislada del paciente con queralitis, una afección que, si bien es de baja prevalencia y su diagnóstico etiológico es de fácil ejecución y bajo costo, no justifica no realizar el estudio oportuna y rápidamente. La sensibilidad de la observación directa por microscopia óptica fue del 11.43%, comparada con los cultivos. Finalmente efectuamos un listado de recomendaciones para usuarios de lentes de contacto.

Evaluation of Three Different Methods to Establish Animal Models of Acanthamoeba Keratitis

PURPOSE: To produce animal models of Acanthamoeba keratitis and to evaluate the advantages and adaptation range of each of the three methods employed. MATERIALS AND METHODS: Mice and Wistar rats in three groups of 15 rats and 15 mice each were used to establish the models. Right corneas in group A were scratched and challenged with Acanthamoeba. Those in group B were scratched and covered with contact lenses incubated with Acanthamoeba. Those in group C received an intrastromal injection of Acanthamoeba. Five rats and 5 mice in each group were used for histopathological investigations and the other 10 in each group were used for clinical evaluation. The models were evaluated by slit lamp examination, microscopic examination and culture of corneal scrapings, HE staining of corneal sections, and pathological scoring of the infections. RESULTS: Four rats and 6 mice in group A, 7 rats and 8 mice in group B, and 10 rats and 10 mice in group C developed typical Acanthamoeba keratitis. CONCLUSION: Corneal scratching alone has the lowest infection rate, while scratching and then covering with contaminated contact lenses has a moderate rate of infection and most closely mimics what happens in most human infections. Intrastromal injection of Acanthamoeba gives a much higher infection rate and more severe Acanthamoeba keratitis.

Acanthamoeba adherence to soft contact lens and human corneal stroma.

In the present study, the adhesibility of Acanthamoeba castellani trophozoites and cysts to hydrogel contact lenses and to human cornea was investigated. Segments of unworn contact lenses were prepared (120 segments in total). In addition, 8 corneal buttons were obtained from 8 enucleated eyes. And each cornea cut into 8 radial segments. To these lens and corneal segments, 1.2 x 106/ml and 1.3 x 106/ml of cysts and trophozoites respectively were added under varying conditions. The adhesion was checked at 0, 3 and 24 hours after the exposure. Adhesion analysis showed that the trophozoites adhered equally well to lens or cornea. There is an increase in the number of trophozoites adhering to contact lenses as well as cornea. This difference is more significant for contact lenses. Washing of contact lenses significantly decreased the adherence of the trophozoites after 3 hours of incubation. When the comparison of adhesion was done between the unwashed worn and unwashed unworn contact lenses, it was observed that there was a significant difference in adherence to new lenses where the adherence was much lower. The study shows that washing of contact lenses does decrease the chances of colonization by Acantamoeba catellani and also that older lenses have higher chances of getting colonized probably due to the occurrence of scratches et. on the surface which may help in colonization.

Random amplified polymorphic DNA profiles as a tool for the characterization of Brazilian keratitis isolates of the genus Acanthamoeba

The genus Acanthamoeba comprises free-living amebae identified as opportunistic pathogens of humans and other animal species. Morphological, biochemical and molecular approaches have shown wide genetic diversity within the genus. In an attempt to determine the genetic relatedness among isolates of Acanthamoeba we analyzed randomly amplified polymorphic DNA (RAPD) profiles of 11 Brazilian isolates from cases of human keratitis and 8 American type culture collection (ATCC) reference strains. We found that ATCC strains belonging to the same species present polymorphic RAPD profiles whereas strains of different species show very similar profiles. Although most Brazilian isolates could not be assigned with certainty to any of the reference species, they could be clustered according to pattern similarities. The results show that RAPD analysis is a useful tool for the rapid characterization of new isolates and the assessment of genetic relatedness of Acanthamoeba spp. A comparison between RAPD analyses and morphological characteristics of cyst stages is also discussed (au)

Amebas patógenas de vida libre / Pathogenic free living amebas

Las pequeñas amebas de vida libre están ampliamente distribuidas en todo el mundo en continuo contacto con el hombre y animales; sus formas quísticas son capaces de sobrevivir en el suelo, aire y agua. Las infecciones causadas por las mismas han tomado en los últimos treinta años notable importancia médica ya que muchos casos fatales no fueron diagnosticados clínicamente ni por el laboratorio, debido al desconocimiento de la potencial capacidad patogénica de estas amebas. Hoy se sabe que la meningoencefalitis amebiana primaria (MAP) causada por Naegleria fowleri y la encefalitits amebiana granulomatosa (EAG) originada por especies de Acanthamoeba spp se han incrementado en el mundo tanto en sujetos sanos como en inmunocomprometidos, incluyendo muchos individuos con SIDA. El grupo más reciente de infecciones causadas por especies del género Acanthamoeba es la queratitis amebiana relacionada principalmente con la falta de cuidado en el mantenimiento de las lentes de contacto. La terapia de la queratitis es problemática debido a la presencia de quistes en los tejidos, y aunque se han informado algunas curas de pacientes, la terapéutica médica aún constituye un capítulo no resuelto