The family “Marseilleviridae” is a new proposed taxon for giant viruses that infect amoebae. Its first member, Acanthamoeba polyphaga marseillevirus (APMaV), was isolated in 2007 by culturing on amoebae a water sample collected from a cooling tower in Paris, France. APMaV has an icosahedral shape with a diameter of ≈250 nm. Its genome is a double-stranded circular DNA that is 368,454 base pairs (bp) in length. The genome has a GC content of 44.7 % and is predicted to encode 457 proteins. Phylogenetic reconstructions showed that APMaV belongs to a new viral family among nucleocytoplasmic large DNA viruses, a group of viruses that also includes Acanthamoeba polyphaga mimivirus (APMV) and the other members of the family Mimiviridae as well as the members of the families Poxviridae, Phycodnaviridae, Iridoviridae, Ascoviridae, and Asfarviridae. In 2011, Acanthamoeba castellanii lausannevirus (ACLaV), another close relative of APMaV, was isolated from river water in France. The ACLaV genome is 346,754 bp in size and encodes 450 genes, among which 320 have an APMaV protein as the closest homolog. Two other giant viruses closely related to APMaV and ACLaV have been recovered in our laboratory from a freshwater sample and a human stool sample using an amoebal co-culture method. The only currently identified hosts for “marseilleviruses” are Acanthamoeba spp. The prevalence of these viruses in the environment and in animals and humans remains to be determined.
Free-living amoebae (FLA) are potential reservoirs of Legionella in aquatic environments. However, the parasitic relationship between various Legionella and amoebae remains unclear. In this study, surface water samples were gathered from two rivers for evaluating parasitic Legionella. Warmer water temperature is critical to the existence of Legionella. This result suggests that amoebae may be helpful in maintaining Legionella in natural environments because warmer temperatures could enhance parasitisation of Legionella in amoebae. We next used immunomagnetic separation (IMS) to identify extracellular Legionella and remove most free Legionella before detecting the parasitic ones in selectively enriched amoebae. Legionella pneumophila was detected in all the approaches, confirming that the pathogen is a facultative amoebae parasite. By contrast, two obligate amoebae parasites, Legionella-like amoebal pathogens (LLAPs) 8 and 9, were detected only in enriched amoebae. However, several uncultured Legionella were detected only in the extracellular samples. Because the presence of potential hosts, namely Vermamoeba vermiformis, Acanthamoeba spp. and Naegleria gruberi, was confirmed in the samples that contained intracellular Legionella, uncultured Legionella may survive independently of amoebae. Immunomagnetic separation and amoebae enrichment may have referential value for detecting parasitic Legionella in surface waters.
Free-living amoebae of the genus Acanthamoeba are causal agents of a severe sight-threatening infection of the cornea known as Acanthamoeba keratitis. Moreover, the number of reported cases worldwide is increasing year after year, mostly in contact lens wearers, although cases have also been reported in non-contact lens wearers. Interestingly, Acanthamoeba keratitis has remained significant, despite our advances in antimicrobial chemotherapy and supportive care. In part, this is due to an incomplete understanding of the pathogenesis and pathophysiology of the disease, diagnostic delays and problems associated with chemotherapeutic interventions. In view of the devastating nature of this disease, here we present our current understanding of Acanthamoeba keratitis and molecular mechanisms associated with the disease, as well as virulence traits of Acanthamoeba that may be potential targets for improved diagnosis, therapeutic interventions and/or for the development of preventative measures. Novel molecular approaches such as proteomics, RNAi and a consensus in the diagnostic approaches for a suspected case of Acanthamoeba keratitis are proposed and reviewed based on data which have been compiled after years of working on this amoebic organism using many different techniques and listening to many experts in this field at conferences, workshops and international meetings. Altogether, this review may serve as the milestone for developing an effective solution for the prevention, control and treatment of Acanthamoeba infections.
Acanthamoeba keratitis (AK) is a painful and potentially blinding corneal infection caused by Acanthamoeba spp. In Madrid, environmental studies have demonstrated a high presence of these free-living amoebae in tap water. Since most of AK cases occur in contact lenses (CL) wearers with inadequate hygiene habits, the presence of Acanthamoeba in discarded CL has been studied and compared with other common etiological agents of keratitis, such as Pseudomonas aeruginosa and Staphylococcus aureus.
The soil amoebae Acanthamoeba causes Acanthamoeba keratitis, a severe sight-threatening infection of the eye and the almost universally fatal granulomatous amoebic encephalitis. More effective treatments are required. Sterol biosynthesis has been effectively targeted in numerous fungi using azole compounds that inhibit the cytochrome P450 enzyme sterol 14α-demethylase. Herein, using Gas Chromatography Mass Spectrometry (GCMS), we demonstrate that the major sterol of Acanthamoeba castellanii is ergosterol and identify novel putative precursors and intermediate sterols in its production. Unlike previously reported, we find no evidence of 7-dehydrostigmasterol or any other phytosterol in Acanthamoeba. Of five azoles tested, we demonstrate that tioconazole and voriconazole have the greatest overall inhibition for all isolates of Acanthamoeba castellanii and Acanthamoeba polyphaga tested. While miconazole and sulconazole have intermediate activity econazole is least effective. Through GCMS, we demonstrate that voriconazole inhibits 14α-demethylase as treatment inhibits the production of ergosterol, but results in the accumulation of the lanosterol substrate. These data provide the most complete description of sterol metabolism in Acanthamoeba, provide a putative framework for their further study and validate 14α-demethylase as the target for azoles in Acanthamoeba.
Acanthamoebidae is a “family” level amoebozoan group composed of the genera Acanthamoeba, Protacanthamoeba, and very recently Luapeleamoeba. This clade of amoebozoans has received considerable attention from the broader scientific community as Acanthamoeba spp. represent both model organisms and human pathogens. While the classical composition of the group (Acanthamoeba + Protacanthamoeba) has been well accepted due to the morphological and ultrastructural similarities of its members, the Acanthamoebidae has never been highly statistically supported in single gene phylogenetic reconstructions of Amoebozoa either by maximum likelihood (ML) or Bayesian analyses.
The aim was to describe a geographically and clinically diverse sample of cases of Acanthamoeba keratitis (AK) and establish the risk factors for poor outcomes among patients with this disease.
Acanthamoeba species are ubiquitous free-living protozoa that can be found worldwide. Occasionally, it can become parasitic and the causative agent of acanthamoebic keratitis (AK) and Granulomatous Amoebic Encephalitis (GAE) in man. A total of 160 environmental samples and 225 naturally-infected animal corneal swabs were collected for Acanthamoeba cultivation. Acanthamoeba was found to be high in samples collected from environments (85%, 136/160) compared to infected animal corneas (24.89%, 56/225) by microscopic examination. Analysis of nucleotide sequence of 18S rRNA gene of all the 192 cultivable Acanthamoeba isolates revealed 4 genotypes (T3, T4. T5 and T15) with T4 as the most prevalent (69.27%, 133/192) followed by T5 (20.31%), T15 (9.90%) and T3 (0.52%). Genotype T4 was from the strain of A. castellanii U07401 (44.27%), A. castellanii U07409 (20.83%) and A. polyphagaAY026243 (4.17%), but interestingly, only A. castellanii U07401 was detected in naturally infected corneal samples. In environmental samples, T4 was commonly detected in all samples including dry soil, dust, wet debris, wet soil and water. Among the T4, A. castellanii (U07409) strains were detected high occurrence in dry (45%) followed by aquatic (32.50%) and moist (22.50%) samples but however A. castellanii (U07401) strains were dominant in dry samples of soil and dust (93.10%). Subsequently, genotype T5 of A. lenticulata (U94741) strains were dominant in samples collected from aquatic environments (58.97%). In summary, A. castellanii (U07401) strains were found dominant in both environmental and corneal swab samples. Therefore, these strains are possibly the most virulent and dry soil or dusts are the most possible source of Acanthamoeba infection in cats and dogs corneas.
A high diversity of microorganisms is encountered inside dental unit waterlines (DUWL). Among those the presence of free-living amoebae (FLA) appears currently underestimated, although human infections may occur due to contact with FLA-contaminated water during dental cares. In order to limit microbial DUWL contamination, disinfectants are provided by dental unit manufacturer, however, with limited documentation on their activities against FLA. The aim of this study was to evaluate the efficiency of three commercial DUWL disinfectants: the Calbenium© (Airel, Champigny-sur-Marne, France), the Oxygenal 6© (Kavo, Biberach, Germany) and the Sterispray© (Gammasonic, Billom, France), against two FLA species, i.e. Acanthamoeba castellanii and Vermamoeba vermiformis alone or co-cultured with Pseudomonas aeruginosa and Candida albicans at concentrations ranging from 0% to 5% (v/v). Results showed varied efficacies of disinfectants: the Oxygenal 6© did not exhibit FLA killing activity, while the Sterispray© and the Calbenium© displayed concentration- and species-dependent activities with a maximum eradication rates of 100% and 86%, and 79% and 97% for A. castellani and V. vermiformis, respectively. None of the disinfectants were able to totally eradicate FLA at concentrations recommended by manufacturers. Present results highlight unsatisfactory anti-FLA activities of 3 DUWL disinfectant preparations advocating deeper investigation of antimicrobial spectra of commercial disinfectants in use for DUWL maintenance.