Expression of matrix metalloproteinases in Naegleria fowleri and their role in invasion of the central nervous system.

Naegleria fowleri is a free-living amoeba found in freshwater lakes and ponds and is the causative agent of primary amoebic meningoencephalitis (PAM), a rapidly fatal disease of the central nervous system (CNS). PAM occurs when amoebae attach to the nasal epithelium and invade the CNS, a process that involves binding to, and degradation of, extracellular matrix (ECM) components. This degradation is mediated by matrix metalloproteinases (MMPs), enzymes that have been described in other pathogenic protozoa, and that have been linked to their increased motility and invasive capability. These enzymes also are upregulated in tumorigenic cells and have been implicated in metastasis of certain tumours. In the present study, in vitro experiments linked MMPs functionally to the degradation of the ECM. Gelatin zymography demonstrated enzyme activity in N. fowleri whole cell lysates, conditioned media and media collected from invasion assays. Western immunoblotting indicated the presence of the metalloproteinases MMP-2 (gelatinase A), MMP-9 (gelatinase B) and MMP-14 [membrane type-1 matrix metalloproteinase (MT1-MMP)]. Highly virulent mouse-passaged amoebae expressed higher levels of MMPs than weakly virulent axenically grown amoebae. The functional relevance of MMPs in media was indicated through the use of the MMP inhibitor, 1,10-phenanthroline. The collective in vitro results suggest that MMPs play a critical role in vivo in invasion of the CNS and that these enzymes may be amenable targets for limiting PAM.

Acanthamoeba T4, T5 and T11 isolated from mineral water bottles in southern Brazil.

Acanthamoeba is a protist potential pathogen, capable of causing a blinding keratitis in contact lens wearers and disseminated infection, leading to granulomatous amebic encephalitis in immunocompromised individuals. This amoeba is a ubiquitous organism that has been isolated from various domestic water systems, such as cooling towers and hospital water networks. The objective of this work was to investigate the presence of Acanthamoeba in mineral water bottles marketed in Porto Alegre, southern Brazil. Positive samples were further classified at the genotype level after sequencing the ASA.S1 region of 18S rDNA gene. Six of the eight isolates belonged to T5 genotype, one to T4 genotype, and one was T11. Several genotypes have been reported worldwide as causative of pathologies in humans, including genotypes T4, T5 and T11. Overall, the widespread distribution of potentially pathogenic Acanthamoeba strains in the studied source demands more awareness within the public and health professionals, because this pathogen is emerging as a risk for human health worldwide.

Prevalence of Acanthamoeba spp. (Sarcomastigophora: Acanthamoebidae) in wild populations of Aedes aegypti (Diptera: Culicidae).

Studies of interrelationship between microorganisms and mosquitoes are of great importance, since it can provide support for better understand related to biology, development and their control. In this way, it is known that mosquito larvae and free-living amoebae (FLA) normally occupy similar aquatic microhabitats. However, few studies have been conducted about such coexistence. For that reason, the objective of the present study was to verify the prevalence of Acanthamoeba spp. in wild populations of Aedes aegypti, as well as to characterize the genotypic lineage, and their possible pathogenicity through thermo- and osmotolerance. Amoebae were investigated in 60 pools, each containing ten larvae of A. aegypti, collected in Porto Alegre (Rio Grande do Sul, Brazil). The Acanthamoeba isolates were morphologically characterized and submitted to the polymerase chain reaction technique to confirm identification of the genus. In addition, genotype analyses as well as tests for presumptive pathogenicity in some samples were performed. Of the 60 pools examined, 54 (90 %) were positive for FLA. Of these isolates, 47 (87 %) belonged to the genus Acanthamoeba. The genotypic groups T4, T3 and T5 were identified, numbering 14 (53.8 %), ten (38.5 %) and two (7.7 %) isolates, respectively. The physiological tests performed with 14 strains showed that 12 (85.7 %) were non-pathogenic, while two (14.3 %) were considered as having low pathogenic potential. These results provide a basis for a better understanding of the interaction between these protozoan and mosquitoes in their natural habitat. This study is the first to report the isolation of Acanthamoeba spp. from wild mosquitoes.

Potentially pathogenic acanthamoeba isolated from a hospital in Brazil.

Studies on free-living amoebae (FLA), has been increased in recent years, especially related to the genus Acanthamoeba, because these organisms are widely found in the environment. The present work isolated and characterized this organism from biofilms and dust in hospital environment. 135 samples were collected in 15 different environments in a hospital at the south of Brazil. Thirty-one (23%) isolates were identified as morphologically belonging to the Acanthamoeba genus and 10 of these were submitted to temperature and osmotolerance tests as criterion for evaluation of the viability and pathogenicity. The tests indicate that four (40%) of these isolates could be potentially pathogenic because grew at high temperature (40 degrees C) and osmolarity (mannitol 1 M). Some isolates genotypes were determined after ribosomal DNA sequencing. These data revealed that three dust isolates belong to T4, two biofilm isolates to T5 and one dust isolate to T3 genotype. Therefore, Acanthamoeba found in the hospital environment represents a risk for people that circulate there.

[Isolation and identification of potentially pathogenic free-living amoebae in samples from environments in a public hospital in the city of Porto Alegre, Rio Grande do Sul]. / Isolamento e identificação de amebas de vida livre potencialmente patogênicas em amostras de ambientes de hospital público da cidade de Porto Alegre, RS.

A study on the presence of free-living amoebae in a public hospital was developed in the city of Porto Alegre, State of Rio Grande do Sul. Dust and biofilms were collected using sterile swabs that had been prepared for this study, from 15 hospital environments, including the intensive care center, pediatric intensive care unit, kitchen, emergency room, outpatient surgical center, clinical surgical center, water storage tanks, taps and six drinking fountains for general use, every month from July 2004 to March 2005. The FLAs were isolated by culturing, using non-nutrient agar medium with the addition of heat-killed Escherichia coli. The protozoa were identified by morphological observation of cysts and trophozoites, in accordance with Page's morphological criteria (1988). Among the 135 samples collected from the 15 environments, 47 (35%) were positive for FLAs. Of these, thirty-four percent presented morphological characteristics particular to the genus Acanthamoeba.

Isolamento e identificação de amebas de vida livre potencialmente patogênicas em amostras de ambientes de hospital público da cidade de Porto Alegre, RS / Isolation and identification of potentially pathogenic free-living amoebae in samples from environments in a public hospital in the city of Porto Alegre, Rio Grande do Sul

Um estudo sobre a presença de amebas de vida livre em um hospital público foi desenvolvido na Cidade de Porto Alegre, RS. Poeira e biofilmes de 15 ambientes hospitalares, incluindo CTI, UTI pediátrica, cozinha, emergência, centro cirúrgico ambulatorial e centro cirúrgico, reservatórios de água, torneira e 6 bebedouros coletivos foram coletados mensalmente, de julho de 2004 a março de 2005, usando-se suabes estéreis, preparados para a pesquisa. As AVL foram isoladas em cultivo, utilizando-se meio de ágar não nutriente adicionado de Escherichia coli, mortas pelo calor. A identificação dos protozoários foi feita pela observação morfológica de cistos e trofozoítos, segundo critérios morfológicos de Page (1988). Das 135 amostras coletadas dos 15 ambientes estudados, 47 (35 por cento) foram positivas para AVL. Destas, 34 por cento apresentaram características morfológicas próprias do gênero Acanthamoeba.

A study on the presence of free-living amoebae in a public hospital was developed in the city of Porto Alegre, State of Rio Grande do Sul. Dust and biofilms were collected using sterile swabs that had been prepared for this study, from 15 hospital environments, including the intensive care center, pediatric intensive care unit, kitchen, emergency room, outpatient surgical center, clinical surgical center, water storage tanks, taps and six drinking fountains for general use, every month from July 2004 to March 2005. The FLAs were isolated by culturing, using non-nutrient agar medium with the addition of heat-killed Escherichia coli. The protozoa were identified by morphological observation of cysts and trophozoites, in accordance with Page's morphological criteria (1988). Among the 135 samples collected from the 15 environments, 47 (35 percent) were positive for FLAs. Of these, thirty-four percent presented morphological characteristics particular to the genus Acanthamoeba.