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Pierre Pernin - One of the best experts on this subject based on the ideXlab platform.
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use of multiplex pcr and pcr restriction enzyme analysis for detection and exploration of the variability in the free living amoeba Naegleria in the environment
Applied and Environmental Microbiology, 2002Co-Authors: Michel Pelandakis, Pierre PerninAbstract:A multiplex PCR was developed to simultaneously detect Naegleria fowleri and other Naegleria species in the environment. Multiplex PCR was also capable of identifying N. fowleri isolates with internal transcribed spacers of different sizes. In addition, restriction fragment length polymorphism analysis of the PCR product distinguished the main thermophilic Naegleria species from the sampling sites.
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analysis of the 5 8s rrna gene and the internal transcribed spacers in Naegleria spp and in n fowleri
Journal of Eukaryotic Microbiology, 2000Co-Authors: Michel Pelandakis, Stephanie Serre, Pierre PerninAbstract:Abstract Internal transcribed spacers (ITS) and the 5.8S ribosomal gene of 21 Naegleria fowleri strains and eight other species including Naegleria gruberi were sequenced. The results showed that this region can help differentiate between and within species. The phylogeny of Naegleria spp. deduced from the ITS and the 5.8S gene produced four major lineages, fowleri-lovaniensis, galeacystis-italica-clarki-gruberi-australiensis, andersoni-jamiesoni, and pussardi, that fit perfectly with those inferred from the 18S rRNA gene analysis. The N. gruberi isolate, NG260, was closely related to Naegleria pussardi. The other N. gruberi isolates branched together with Naegleria australiensis in another lineage. The ITS and 5.8S results for N. fowleri were congruent with those previously deduced by RAPD analysis. The phylogenetic analysis inferred from ITS and RAPD data revealed two major groups. The French Cattenom and Chooz and South Pacific strains constituted the first group. The second group encompassed the strai...
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analysis of the 5 8s rrna gene and the internal transcribed spacers in Naegleria spp and in n fowleri
Journal of Eukaryotic Microbiology, 2000Co-Authors: Michel Pelandakis, Stephanie Serre, Pierre PerninAbstract:Internal transcribed spacers (ITS) and the 5.8S ribosomal gene of 21 Naegleria fowleri strains and eight other species including Naegleria gruberi were sequenced. The results showed that this region can help differentiate between and within species. The phylogeny of Naegleria spp. deduced from the ITS and the 5.8S gene produced four major lineages, fowleri-lovaniensis, galeacystis-italica-clarki-gruberi-australiensis, andersoni-jamiesoni, and pussardi, that fit perfectly with those inferred from the 18S rRNA gene analysis. The N. gruberi isolate, NG260, was closely related to Naegleria pussardi. The other N. gruberi isolates branched together with Naegleria australiensis in another lineage. The ITS and 5.8S results for N. fowleri were congruent with those previously deduced by RAPD analysis. The phylogenetic analysis inferred from ITS and RAPD data revealed two major groups. The French Cattenom and Chooz and South Pacific strains constituted the first group. The second group encompassed the strains corresponding to the Euro-American and Widespread RAPD variants and shared the same substitution in the 5.8S gene. In addition, it was possible to define species specific primers in ITS regions to rapidly identify N. fowleri.
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comparative recoveries of Naegleria fowleri amoebae from seeded river water by filtration and centrifugation
Applied and Environmental Microbiology, 1998Co-Authors: Pierre Pernin, Michel Pelandakis, Y Rouby, A Faure, F SicletAbstract:Detection of pathogenic Naegleria fowleri in environmental water samples, which is necessary for the prevention of primary amoebic meningoencephalitis, generally requires concentrating the samples. Two concentration techniques, filtration and centrifugation, were used to study the recovery of N. fowleri, in vegetative or cystic form, that had been mixed with the two other thermotolerant Naegleria species, N. lovaniensis and N. australiensis. Counting of amoebae was performed by the most probable number method on 10 water replicates of 100 ml and 10 ml each. With both concentration methods, recovery was better for cysts than for trophozoites (53% ± 21% versus 5% ± 5% by filtration and 57% ± 25% versus 22% ± 5% by centrifugation). The recovery of Naegleria trophozoites by filtration was very low, and centrifugation was significantly better than filtration in recovery of Naegleria trophozoites (22% ± 5% versus 5% ± 5%; P 0.7). Although the recovery of cysts of N. fowleri obtained by filtration (51% ± 24%) appeared higher than that by centrifugation (36% ± 23%), the difference was not significant (P > 0.1). Both concentration methods have highly variable recovery rates, making accurate quantification of low concentrations (<100/liter) of N. fowleri in the environment difficult.
Bingmu Hsu - One of the best experts on this subject based on the ideXlab platform.
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Application of TaqMan qPCR for the detection and monitoring of Naegleria species in reservoirs used as a source for drinking water
Parasitology Research, 2014Co-Authors: Po-min Kao, Bingmu Hsu, Y.-c. Chiu, Tsui-kang Hsu, Chung-liang Chang, Shih-wei Huang, Cheng-wei FanAbstract:Naegleria spp. can be found in the natural aquatic environments. Naegleria fowleri can cause fatal infections in the central nervous system in humans and animals, and the most important source of infection is through direct water contact. In this study, PCR of 5.8S ribosomal RNA (rRNA) gene and internal transcribed spacer (ITS) region was performed in order to identify Naegleria isolates and quantify the Naegleria spp. by TaqMan real-time quantitative PCR in reservoir water samples. The occurrence of Naegleria spp. was investigated in 57 water samples from reservoirs with culture and PCR positive in 2 of them (3.5 %), respectively. The total detection rate was 7.0 % (4/ 57) for Naegleria spp. The identified species included Naegleria spp., Naegleria canariensis , and Naegleria clarki. N. fowleri was not found in Taiwan’s reservoirs used for drinking purposes. The concentrations of Naegleria spp. in detected positive reservoir water samples were in the range of 599 and 3.1 × 10^3 cells/L. The presence or absence of Naegleria spp. within the reservoir water samples showed significant difference with the levels of water temperature. The presence of Naegleria spp. in reservoirs considered a potential public health threat if pathogenic species exist in reservoirs.
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Quantitative detection and identification of Naegleria spp. in various environmental water samples using real-time quantitative PCR assay
Parasitology research, 2013Co-Authors: Po-min Kao, Bingmu Hsu, M.-c. Tung, Ming-yuan Chou, Hsiu-wu Yang, Cheng-yu She, Shu-min ShenAbstract:Naegleria spp. is a free-living amoeba that can be found in various aquatic environments. There are some Naegleria spp. that can cause fatal infections in animals and humans, and the most important source of infection is through direct water contact. In this study, a real-time quantitative PCR was developed to detect and quantify the Naegleria spp. in various environmental water samples. The water samples were taken from rivershed, water treatment plants, and thermal spring recreation areas. The total detection rate was 4.0 % (7/176) for Naegleria spp. The percentages of samples containing Naegleria spp. from river water, raw drinking water, and thermal spring water were 0 % (0/100), 10.7 % (3/28) and 8.3 % (4/48), respectively. The concentration of Naegleria spp. in detected positive raw drinking water and thermal spring water samples was in the range of 3.9–12.6 and 1.1–24.2 cells/L, respectively. The identified species included Naegleria australiensis, Naegleria lovaniensis, and Naegleria spitzbergeniensis. The presence of Naegleria spp. in various aquatic environments is considered a potential public health threat.
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Occurrence and distribution of Naegleria species from thermal spring environments in Taiwan.
Letters in applied microbiology, 2012Co-Authors: Po-min Kao, Bingmu Hsu, M.-c. Tung, C.-j. Hsueh, Y.-c. Chiu, Nai-hsiung Chen, Shu-min Shen, Yu-li HuangAbstract:UNLABELLED Naegleria spp. is a free-living amoeba that can be found in the natural environment. A number of Naegleria spp. can cause fatal infections in the central nervous system in humans and animals, and the most important source of infection is through direct water contact. In this study, water samples from various thermal springs were taken from four thermal spring areas. Naegleria spp. was detected via culture confirmation and molecular taxonomic identification. Among the 60 samples obtained, Naegleria spp. was identified in 26 (43·3%) samples. The identified species included Naegleria australiensis, Naegleria gruberi, Naegleria lovaniensis and Naegleria mexicana. The presence of living Naegleria spp. was significantly associated with elevated pH value in the water sample. SIGNIFICANCE AND IMPACT OF STUDY In this study, we examined the presence of living Naegleria spp. in thermal spring waters in south-eastern Taiwan. Naegleria spp. was isolated and culture-confirmed from thermal spring water. Naegleria fowleri was not found in all water samples, and Naegleria australiensis was the most common Naegleria genotype.
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Identification of the Naegleria Species in Natural Watersheds Used for Drinking and Recreational Purposes in Taiwan
Journal of Environmental Engineering, 2012Co-Authors: Po-min Kao, Bingmu Hsu, Y.-c. Chiu, Nai-hsiung Chen, Kuan-hao Huang, Shu-min ShenAbstract:AbstractNaegleria species (spp.) is a free-living amoeboflagellate that is widespread in natural environments. Some Naegleria spp. cause fatal human infections, and the most important source of these infections is water. This genus is recognized as a potential human pathogen that causes primary amoebic meningoencephalitis. In this study, 211 water samples were collected from two watersheds in Taiwan. The Naegleria spp. on the basis of polymerase chain reaction (PCR) amplification with a genus-specific primer pair and an investigation of Naegleria spp. in the Puzih Rivers and Kaoping Rivers in Taiwan. The percentage of positive samples obtained from the Kaoping River (21.8%) was higher than that from the Puzih River (0.6%). The presence/absence of Naegleria spp. within the water samples showed a significant difference with levels of water temperature and pH value. The most frequently identified Naegleria spp. was N. philippinensis (n=3), followed by N. clarki (n=2), N. gallica (n=2), and N. americana (n=2)...
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Survey of Naegleria from Taiwan recreational waters using culture enrichment combined with PCR.
Acta tropica, 2011Co-Authors: Shih-wei Huang, Bingmu HsuAbstract:Naegleria is a free-living amoeba. Pathogenic Naegleria may pose a health risk to people who come in contact with recreational waters. Here, we used Naegleria culture enrichment with PCR to identify the Naegleria species and investigated the distribution of Naegleria spp. in recreational waters including spring water, stream water and raw domestic water in central and southern Taiwan. In this study, Naegleria spp. were detected in 19 (17.8%) of the water samples. The occurrence of Naegleria in raw domestic water was 28.6%, higher than in stream water (14.7%) and in spring water (6.5%). The most frequently identified species exhibiting the closest phylogenetic relationships to the isolates were N. australiensis (n=4) and N. canariensis (n=4), followed by N. clarki (n=3) and N. philippinensis (n=3); N. americana (n=2). N. lovaniensis, N. dobsoni, and N. gruberi were each detected once. The pathogenic species N. fowleri was not detected, probably due to the low incubation temperature; however, the isolates exhibiting the closest phylogenetic relationships to the pathogenic species in mice of PAM, N. australiensis and N. philippinensis, were found. Results of this survey suggest the distribution of Naegleria spp. excluding N. fowleri in recreational waters. It should be considered a potential threat for health associated with human activities in recreational waters.
Michel Pelandakis - One of the best experts on this subject based on the ideXlab platform.
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use of multiplex pcr and pcr restriction enzyme analysis for detection and exploration of the variability in the free living amoeba Naegleria in the environment
Applied and Environmental Microbiology, 2002Co-Authors: Michel Pelandakis, Pierre PerninAbstract:A multiplex PCR was developed to simultaneously detect Naegleria fowleri and other Naegleria species in the environment. Multiplex PCR was also capable of identifying N. fowleri isolates with internal transcribed spacers of different sizes. In addition, restriction fragment length polymorphism analysis of the PCR product distinguished the main thermophilic Naegleria species from the sampling sites.
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analysis of the 5 8s rrna gene and the internal transcribed spacers in Naegleria spp and in n fowleri
Journal of Eukaryotic Microbiology, 2000Co-Authors: Michel Pelandakis, Stephanie Serre, Pierre PerninAbstract:Abstract Internal transcribed spacers (ITS) and the 5.8S ribosomal gene of 21 Naegleria fowleri strains and eight other species including Naegleria gruberi were sequenced. The results showed that this region can help differentiate between and within species. The phylogeny of Naegleria spp. deduced from the ITS and the 5.8S gene produced four major lineages, fowleri-lovaniensis, galeacystis-italica-clarki-gruberi-australiensis, andersoni-jamiesoni, and pussardi, that fit perfectly with those inferred from the 18S rRNA gene analysis. The N. gruberi isolate, NG260, was closely related to Naegleria pussardi. The other N. gruberi isolates branched together with Naegleria australiensis in another lineage. The ITS and 5.8S results for N. fowleri were congruent with those previously deduced by RAPD analysis. The phylogenetic analysis inferred from ITS and RAPD data revealed two major groups. The French Cattenom and Chooz and South Pacific strains constituted the first group. The second group encompassed the strai...
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analysis of the 5 8s rrna gene and the internal transcribed spacers in Naegleria spp and in n fowleri
Journal of Eukaryotic Microbiology, 2000Co-Authors: Michel Pelandakis, Stephanie Serre, Pierre PerninAbstract:Internal transcribed spacers (ITS) and the 5.8S ribosomal gene of 21 Naegleria fowleri strains and eight other species including Naegleria gruberi were sequenced. The results showed that this region can help differentiate between and within species. The phylogeny of Naegleria spp. deduced from the ITS and the 5.8S gene produced four major lineages, fowleri-lovaniensis, galeacystis-italica-clarki-gruberi-australiensis, andersoni-jamiesoni, and pussardi, that fit perfectly with those inferred from the 18S rRNA gene analysis. The N. gruberi isolate, NG260, was closely related to Naegleria pussardi. The other N. gruberi isolates branched together with Naegleria australiensis in another lineage. The ITS and 5.8S results for N. fowleri were congruent with those previously deduced by RAPD analysis. The phylogenetic analysis inferred from ITS and RAPD data revealed two major groups. The French Cattenom and Chooz and South Pacific strains constituted the first group. The second group encompassed the strains corresponding to the Euro-American and Widespread RAPD variants and shared the same substitution in the 5.8S gene. In addition, it was possible to define species specific primers in ITS regions to rapidly identify N. fowleri.
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comparative recoveries of Naegleria fowleri amoebae from seeded river water by filtration and centrifugation
Applied and Environmental Microbiology, 1998Co-Authors: Pierre Pernin, Michel Pelandakis, Y Rouby, A Faure, F SicletAbstract:Detection of pathogenic Naegleria fowleri in environmental water samples, which is necessary for the prevention of primary amoebic meningoencephalitis, generally requires concentrating the samples. Two concentration techniques, filtration and centrifugation, were used to study the recovery of N. fowleri, in vegetative or cystic form, that had been mixed with the two other thermotolerant Naegleria species, N. lovaniensis and N. australiensis. Counting of amoebae was performed by the most probable number method on 10 water replicates of 100 ml and 10 ml each. With both concentration methods, recovery was better for cysts than for trophozoites (53% ± 21% versus 5% ± 5% by filtration and 57% ± 25% versus 22% ± 5% by centrifugation). The recovery of Naegleria trophozoites by filtration was very low, and centrifugation was significantly better than filtration in recovery of Naegleria trophozoites (22% ± 5% versus 5% ± 5%; P 0.7). Although the recovery of cysts of N. fowleri obtained by filtration (51% ± 24%) appeared higher than that by centrifugation (36% ± 23%), the difference was not significant (P > 0.1). Both concentration methods have highly variable recovery rates, making accurate quantification of low concentrations (<100/liter) of N. fowleri in the environment difficult.
Johan F. De Jonckheere - One of the best experts on this subject based on the ideXlab platform.
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What do we know by now about the genus Naegleria?
Experimental parasitology, 2014Co-Authors: Johan F. De JonckheereAbstract:In this short overview of the genus Naegleria a brief historical sketch is given since the discovery of this amoeboflagellate in 1899 and the finding in 1970 that one species, Naegleria fowleri causes primary amoebic meningoencephalitis in man. Eight different types of this pathogen are known which have an uneven distribution over the world. Until now 47 different Naegleria spp. are described, of which two other species cause disease in experimental animals, and their geographical dispersal is indicated. The presence of group I introns in the SSU and in the LSU rDNA in the genus is discussed, as well as the possibility of sex or mating. It is also mentioned that the genome of N. fowleri should not be compared to that of Naegleria gruberi, to know why the former is pathogenic, but to the genome of its closest relative Naegleria lovaniensis.
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origin and evolution of the worldwide distributed pathogenic amoeboflagellate Naegleria fowleri
Infection Genetics and Evolution, 2011Co-Authors: Johan F. De JonckheereAbstract:Naegleria fowleri, a worldwide distributed pathogen, is the causative agent of primary amoebic meningoencephalitis. Because it is such a fulminant disease, most patients do not survive the infection. This pathogen is a free-living amoeboflagellate present in warm water. To date, it is well established that there are several types of N. fowleri, which can be distinguished based on the length of the internal transcribed spacer 1 and a one bp transition in the 5.8S rDNA. Seven of the eight known types have been detected in Europe. Three types are present in the USA, of which one is unique to this country. Only one of the eight types occurs in Oceania (Australia and New Zealand) and Japan. In mainland Asia (India, China and Thailand) the two most common types are found, which are also present in Europe and the USA. There is strong indication that the pathogenic N. fowleri evolved from the nonpathogenic Naegleria lovaniensis on the American continent. There is no evidence of virulence differences between the types of N. fowleri. Two other Naegleria spp. are pathogenic for mice, but human infections due to these two other Naegleria spp. are not known.
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Naegleria gruberi metabolism.
International journal for parasitology, 2011Co-Authors: Fred R. Opperdoes, Johan F. De Jonckheere, Aloysius G. M. TielensAbstract:The completion of the genome project for Naegleria gruberi provides a unique insight into the metabolic capacities of an organism, for which there is an almost complete lack of experimental data. The metabolism of Naegleria seems to be extremely versatile, as can be expected for a free-living amoeboflagellate, but although considered to be fully aerobic, its genome also predicts important anaerobic traits. Other predictions are that carbohydrates are oxidised to carbon dioxide and water when oxygen is not limiting and that in the absence of oxygen the end-products will be succinate, acetate and minor quantities of ethanol and D-lactate. The hybrid mitochondrion/hydrogenosome has both cytochromes and an [Fe] hydrogenase, but seems to lack pyruvate-ferredoxin oxidoreductase. Genomic information also provides the possibility to identify drugs with a possible mode of action in the fatal primary amoebic meningoencephalitis caused by the closely related opportunistic pathogen Naegleria fowleri.
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Molecular definition and the ubiquity of species in the genus Naegleria.
Protist, 2004Co-Authors: Johan F. De JonckheereAbstract:Summary To investigate the variability within species of the genus Naegleria , the ITS1, 5.8S and ITS2 rDNA were sequenced of several strains of N. lovaniensis and its Western Australian variants, N. australiensis , N.fowleri , N. andersoni , N. jamiesoni , N. tihangensis , N. pringsheimi , N. pagei , N. gruberi sensu lato and a Naegleria lineage that lost a group I intron from the SSUrDNA twintron. As a result, it is possible to define a molecular species within the Naegleria genus. In addition, one strain of each different allozyme cluster was sequenced to investigate whether they belong to described species or should be treated as distinct new species. This leads to the proposal of eleven new species. The sequencing results from those Naegleria spp. of which several strains are available indicate that these species are ubiquitous. The only exception might be the species represented by the WA variants. However, there are still many Naegleria spp. for which only one strain has been isolated, hence, it is important that the search for more isolates should be continued worldwide.
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Three different group I introns in the nuclear large subunit ribosomal DNA of the amoeboflagellate Naegleria
Nucleic acids research, 1998Co-Authors: Johan F. De Jonckheere, Susan J. BrownAbstract:We have amplified the large subunit ribosomal DNA (LSUrDNA) of the 12 described Naegleria spp. and of 34 other Naegleria lineages that might be distinct species. Two strains yielded a product that is longer than 3 kb, which is the length of the LSUrDNA of all described Naegleria spp. Sequencing data revealed that the insert in one of these strains is a group I intron without an open reading frame (ORF), while the other strain contains two different group I introns, of which the second intron has an ORF of 175 amino acids. In the latter ORF there is a conserved His-Cys box, as in the homing endonucleases present in group I introns in the small subunit ribosomal DNA (SSUrDNA) of Naegleria spp. Although the group I introns in the LSUrDNA differ in sequence, they are more related to each other than they are to the group I introns in the SSUrDNA of Naegleria spp. The three group I introns in the LSUrDNA in Naegleria are at different locations and are probably acquired by horizontal transfer, contrary to the SSUrDNA group I introns in this genus which are of ancestral origin and are transmitted vertically.
Susan J. Brown - One of the best experts on this subject based on the ideXlab platform.
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Three different group I introns in the nuclear large subunit ribosomal DNA of the amoeboflagellate Naegleria
Nucleic acids research, 1998Co-Authors: Johan F. De Jonckheere, Susan J. BrownAbstract:We have amplified the large subunit ribosomal DNA (LSUrDNA) of the 12 described Naegleria spp. and of 34 other Naegleria lineages that might be distinct species. Two strains yielded a product that is longer than 3 kb, which is the length of the LSUrDNA of all described Naegleria spp. Sequencing data revealed that the insert in one of these strains is a group I intron without an open reading frame (ORF), while the other strain contains two different group I introns, of which the second intron has an ORF of 175 amino acids. In the latter ORF there is a conserved His-Cys box, as in the homing endonucleases present in group I introns in the small subunit ribosomal DNA (SSUrDNA) of Naegleria spp. Although the group I introns in the LSUrDNA differ in sequence, they are more related to each other than they are to the group I introns in the SSUrDNA of Naegleria spp. The three group I introns in the LSUrDNA in Naegleria are at different locations and are probably acquired by horizontal transfer, contrary to the SSUrDNA group I introns in this genus which are of ancestral origin and are transmitted vertically.
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Willaertia minor is a species of Naegleria
European Journal of Protistology, 1995Co-Authors: Johan F. De Jonckheere, Susan J. BrownAbstract:Summary Riboprinting, or restriction fragment length polymorphism analysis of the PCR-amplified ribosomal RNA gene, demonstrates that Willaertia minor belongs to the genus Naegleria. The species differs from other Naegleria spp. in the ability to divide in the quadriflagellate stage, although there is evidence that division in the flagellate stage of Naegleria spp. was not recognized as such before. Strain WT043, on which the description of W. minor was based, harbors a DNA plasmid of approx. 6 kb of unknown identity.
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Loss of the ORF in the SSUrDNA group I intron of one Naegleria lineage
Nucleic acids research, 1994Co-Authors: Johan F. De Jonckheere, Susan J. BrownAbstract:We have found a Naegleria lineage in which the SSUrDNA contains a group I intron with a length of 375 nucleotides. This is a unique finding because all group I introns detected until now in Naegleria are 1.3 kilobases long and contain an open reading frame coding for 245 amino acids. Sequence data show that the 375 nucleotide-long intron is at the same place in the SSUrDNA as, and is descendant from, the 1.3 kilobase group I intron present in other species of Naegleria. Our data indicate that in one lineage of Naegleria the group I intron lost part of its DNA that is not contributing to the secondary structure but that carries the open reading frame. The amoeboflagellate genus Naegleria contains strains without the intron and strains with the intron, with or without an open reading frame. Therefore, this genus provides a unique opportunity to study the function and evolution of both the group I intron and the open reading frame.
