Prevalence of trichomonads in the cloaca of wild wetland birds in the Netherlands.

Severe granulomatosis in productive layer chickens due to Tetratrichomonas gallinarum strain 13/16632 infection occurred in 2013 and 2017 on farms situated in a wetland area in the Netherlands. We hypothesized that wetland birds could be the source of the infection. Therefore, a prevalence study on trichomonads was performed by analysing cloaca swabs of 526 birds belonging to 13 species of wetland birds. The number of birds sampled ranged from 1 to 275 per species. Birds were sampled at 15 locations in the Netherlands. DNA extracted from the cloaca swabs was subjected to nested PCR using trichomonad-specific primers targeting the internal transcribed spacer 1 (ITS1)-5.8S rRNA-ITS2 region followed by cloning and sequencing. In nine bird species, trichomonads were detected; the overall prevalence was 9% (47/526), while the prevalence in the five species for which a substantial number of birds were examined (at least 39 per species) ranged from 4% to 24%. Three trichomonad species were found: T. gallinarum, Trichomonas tenax and Simplicimonas sp. of which T. gallinarum dominated. The virulent T. gallinarum strain 13/16632 was not detected, but closely related strains were. Phylogenetic analysis revealed that all T. gallinarum isolates belonged to two clusters within lineage 15 of Tetratrichomonas lineages. All T. tenax isolates were identical and clustered with reference strain H95, while Simplicimonas sp. isolates showed large genetic diversity. Some isolates may represent a new species of the genus Simplicimonas. We conclude that trichomonads are widespread amongst wetland birds, raising the question, amongst others, of their relevance for commercial poultry.RESEARCH HIGHLIGHTSTrichomonads occur among wild wetland birds in the Netherlands.T. gallinarum is the dominant trichomonad species in the cloaca of wetland birds.Some T. gallinarum isolates are closely related to a strain causing granulomas in layer chickens.Some isolates may represent a new species of the genus Simplicimonas.

Tetratrichomonas gallinarum granuloma disease in a flock of free range layers.

Granuloma disease in a flock of free range productive layers in the Netherlands in 2017 is described. The disease resembled granuloma outbreaks in layers caused by Tetratrichomonas gallinarum in 2013 and occurred in the same area in which the rearing farm considered as the source of the 2013 outbreaks was located. Between 55 and 84 weeks of age mortality was 20.3% (breeder's norm 3.9%). All dead hens examined (n = 20) showed granulomas especially in liver and ceca. Nine hens with or without liver and/or ceca granulomas were examined for trichomonads in mentioned organs by in situ hybridization (ISH), nested PCR, and cloning and sequencing. Ceca were also examined by culture. T. gallinarum ISH was positive in all livers and ceca with granulomas and negative in case granulomas were absent. T. gallinarum strain 13/16632, which caused the 2013 outbreaks was found in 4/8 hens with granulomas. Moreover, other trichomonads were detected: a T. gallinarum strain GPO-like and a Simplicimonas sp. strain GABC1-like. Mixed infections also occurred. Infectious causes of granuloma disease other than the afore-mentioned trichomonads could be excluded. Trichomonad DNA was not detected in environmental samples and wild ducks originating from the farm of concern, except for one duck in which the same Simplicimonas sp. as in hens was detected, leaving the source of the T. gallinarum infection in hens unknown. It is concluded that the herein described granuloma disease likely was caused by T. gallinarum strain 13/16632. However, the pathogenicity of the other trichomonads found remains to be clarified.

Prevalence and genetic diversity of Campylobacter spp. in the production chain of broiler chickens in Lebanon and its association with the intestinal protozoan Blastocystis sp.

Campylobacter jejuni is recognized as the most common foodborne pathogen associated with human gastroenteritis worldwide. Broilers are frequently infected by the bacteria and are considered the main source of exposure to humans. However, despite its public health impact, no recent data are currently available in Lebanon about Campylobacter spp. in poultry and human population. Therefore, this study aimed to determine the prevalence and genetic diversity of Campylobacter spp. in 227 ceca and on 227 carcasses of broiler chickens collected in Lebanese slaughterhouses. Overall, the prevalence of Campylobacter was shown to reach 67.0% in ceca and 17.2% on carcasses of Lebanese poultry. The only 2 Campylobacter species identified were C. jejuni and C. coli, with a slightly higher prevalence of C. coli in ceca and of C. jejuni on carcasses. A high level of genetic diversity was reported among the 51 C. jejuni isolates selected, since 25 distinct profiles were identified according to the comparative genomic fingerprinting typing method based on a subset of 40 genes using the 90% similarity threshold. Predominant clusters observed in Lebanese poultry isolates were also frequently found among French human clinical cases, highlighting that broiler chickens represent a potential reservoir for human campylobacteriosis. In addition, a significantly higher prevalence of Campylobacter spp. was found in slaughterhouse workers than in a cohort of hospitalized patients with no contact with poultry, confirming that contaminated broiler chickens in slaughterhouse appeared to be a non-negligible source of Campylobacter spp. transmission. Interestingly, a significant association between Campylobacter spp. and Blastocystis sp. has been observed. This correlation suggested that the presence of Campylobacter spp. would be favored when Blastocystis sp. is present and, similarly, the absence of one would favor the absence of the other. This is the first large-scale investigation focusing on the impact of Campylobacter spp. in broiler chickens in Lebanon and confirmed the need to implement prevention and control measures in the poultry production to reduce the burden of campylobacteriosis in the human population.

Emergence of Aspergillus fumigatus azole resistance in azole-naïve patients with chronic obstructive pulmonary disease and their homes.

Azole-resistant Aspergillus fumigatus (ARAF) has been reported in patients with chronic obstructive pulmonary disease (COPD) but has not been specifically assessed so far. Here, we evaluated ARAF prevalence in azole-naïve COPD patients and their homes, and assessed whether CYP51A mutations were similar in clinical and environmental reservoirs. Sixty respiratory samples from 41 COPD patients with acute exacerbation and environmental samples from 36 of these patient's homes were prospectively collected. A. fumigatus was detected in respiratory samples from 11 of 41 patients (27%) and in 15 of 36 domiciles (42%). Cyp51A sequencing and selection on itraconazole medium of clinical (n = 68) and environmental (n = 48) isolates yielded ARAF detection in 1 of 11 A. fumigatus colonized patients with COPD (9%) and 2 of 15 A. fumigatus-positive patient's homes (13%). The clinical isolate had no CYP51A mutation. Two environmental isolates from two patients harbored TR34 /L98H mutation, and one had an H285Y mutation. Coexistence of different cyp51A genotypes and/or azole resistance profiles was detected in 3 of 8 respiratory and 2 of 10 environmental samples with more than one isolate, confirming the need for a systematic screening of all clinically relevant isolates. The high prevalence of ARAF in patients with COPD and their homes supports the need for further studies to assess the prevalence of azole resistance in patients with Aspergillus diseases in Northern France.

A review of methods for nematode identification.

Nematodes are non-segmented roundworms found in soil, aquatic environment, plants, or animals. Either useful or pathogenic, they greatly influence environmental equilibrium, human and animal health, as well as plant production. Knowledge on their taxonomy and biology are key issues to answer the different challenges associated to these organisms. Nowadays, most of the nematode taxonomy remains unknown or unclear. Several approaches are available for parasite identification, from the traditional morphology-based techniques to the sophisticated high-throughput sequencing technologies. All these techniques have advantages or drawbacks depending on the sample origin and the number of nematodes to be processed. This review proposes an overview of all newly available methods available to identify known and/or unknown nematodes with a specific focus on emerging high-throughput molecular techniques.

On Blastocystis secreted cysteine proteases: a legumain-activated cathepsin B increases paracellular permeability of intestinal Caco-2 cell monolayers.

Blastocystis spp. pathogenic potential remains unclear as these anaerobic parasitic protozoa are frequently isolated from stools of both symptomatic and asymptomatic subjects. In silico analysis of the whole genome sequence of Blastocystis subtype 7 revealed the presence of numerous proteolytic enzymes including cysteine proteases predicted to be secreted. To assess the potential impact of proteases on intestinal cells and gut function, we focused our study on two cysteine proteases, a legumain and a cathepsin B, which were previously identified in Blastocystis subtype 7 culture supernatants. Both cysteine proteases were produced as active recombinant proteins. Activation of the recombinant legumain was shown to be autocatalytic and triggered by acidic pH, whereas proteolytic activity of the recombinant cathepsin B was only recorded after co-incubation with the legumain. We then measured the diffusion of 4-kDa FITC-labelled dextran across Caco-2 cell monolayers following exposition to either Blastocystis culture supernatants or each recombinant protease. Both Blastocystis culture supernatants and recombinant activated cathepsin B induced an increase of Caco-2 cell monolayer permeability, and this effect was significantly inhibited by E-64, a specific cysteine protease inhibitor. Our results suggest that cathepsin B might play a role in pathogenesis of Blastocystis by increasing intestinal cell permeability.

Granuloma disease in flocks of productive layers caused by Tetratrichomonas gallinarum.

In 2013, seven outbreaks of granuloma disease occurred in Dutch flocks of productive layers housed on different farms. These outbreaks were characterized by increased mortality and high incidence of granulomas, mainly in caeca (340/408 hens = 83%) and livers (69/408 hens = 17%). Mortality started to increase between 21 and 35 weeks of age and reached 3.7% to 11.0% exceeding the breeder's norm in periods ranging from 9 to 48 weeks. Some flocks also showed decreased egg production and/or loss of mean egg weight. All affected flocks were linked to one rearing farm, which therefore seemed to be the source of the disease. However, no signs of disease had been observed at this rearing farm. Sentinel hens placed in one of the affected flocks to determine whether the disease had an infectious nature developed granulomas identical to those seen in the outbreaks. Next, by fulfilling Koch's postulates it was shown that Tetratrichomonas gallinarum was the aetiological agent of the granuloma disease. The condition was reproduced in mature specified pathogen free White Leghorn hens (GD - Animal Health, Deventer, the Netherlands) by inoculation via both an artificial and a natural route with a well-defined axenic T. gallinarum isolate obtained from one of the affected flocks. Other causes of granuloma disease were excluded.

Assessment of microscopic and molecular tools for the diagnosis and follow-up of cryptosporidiosis in patients at risk.

Cryptosporidiosis is an important though underreported public health concern. Molecular tools might be helpful in improving its diagnosis. In this study, ZR Fecal DNA MiniPrep™ Kit (ZR) and NucliSens® easyMAG® (EM) were compared using four Cryptosporidium-seeded feces and 29 Cryptosporidium-positive stools. Thereafter, ZR was selected for prospective evaluation of Cryptosporidium detection by 18S rDNA and LAXER quantitative PCR (qPCR) in 69 stools from 56 patients after Cryptosporidium detection by glycerin, modified Ziehl-Neelsen (ZN) and auramine-phenol (AP) stainings. The combination of any of the two extraction methods with 18S qPCR yielded adequate detection of Cryptosporidium in seeded stools, but the ZR kit showed the best performance. All 29 Cryptosporidium-positive samples were positive with 18S qPCR, after both ZR and EM extraction. However, false-negative results were found with LAXER qPCR or nested PCR. Cryptosporidiosis was diagnosed in 7/56 patients. All the microscopic methods enabled the initial diagnosis, but Cryptosporidium was detected in 12, 13, and 14 samples from these seven patients after glycerin, ZN, and AP staining respectively. Among these samples, 14 and 12 were positive with 18S and LAXER qPCR respectively. In two patients, Cryptosporidium DNA loads were found to be correlated with clinical evolution. Although little known, glycerin is a sensitive method for the initial detection of Cryptosporidium. When combined with 18S qPCR, ZR extraction, which had not been evaluated so far for Cryptosporidium, was an accurate tool for detecting Cryptosporidium and estimating the oocyst shedding in the course of infection.

Monitoring of four DNA extraction methods upstream of high-throughput sequencing of Anisakidae nematodes.

Different methods were evaluated to extract DNA from pooled nematodes belonging to Anisakis, Contracaecum, Pseudoterranova and Hysterothylacium genera isolated from edible fish. Pooled DNA extraction is the first and compulsory step to allow the identification of a large number of samples through high-throughput DNA sequencing with drastic time and cost reductions.

Molecular subtyping of Blastocystis spp. using a new rDNA marker from the mitochondria-like organelle genome.

Blastocystis spp. are common anaerobic intestinal protozoa found in both human and animals. They are characterized by a high genetic diversity with at least 17 subtypes (STs) that have been described on the basis of a 600 bp 'barcoding region' from the 18S rDNA gene. However, analysis of the recently sequenced genome of a Blastocystis ST7 isolate (strain B) revealed the presence of multiple variable copies of the 18S rDNA gene, with 17 completely assembled copies. Comparison of the barcoding region from these 17 copies allowed us to classify the 18S rDNA sequences into 6 clusters, each cluster containing identical sequences. Surprisingly, 4 of these clusters had the highest homology with 18S rDNA sequences from 2 other Blastocystis ST7 isolates referred as QQ98-4 and H. These results suggest that the 18S rDNA gene is not the marker of choice to discriminate between strains within STs. In the present study, we identified a single-copy subtyping rDNA marker in the genome of the mitochondria-like organelles (MLOs). Using a partial sequence of the MLO rDNA, we successfully subtyped 66 isolates from both human and animals belonging to Blastocystis ST1 to ST10. Our results also indicate that this mitochondrial marker could be useful to detect co-infections by different isolates of a same ST.

The genotypic characterisation of Acanthamoeba isolates from human ocular samples.

BACKGROUND: We characterised 37 amoebae cultured from corneal scrapings, contact lenses or lens case solutions of patients with suspected Acanthamoeba keratitis. METHODS: The isolates were identified by their morphology and by PCR targeting the Acanthamoeba nuclear small-subunit rRNA gene. Acanthamoeba isolates were genotyped by DNA sequence analysis. RESULTS: The 37 isolates comprised 35 Acanthamoeba, one Hartmannella and one Vahlkampfia. Ten Acanthamoeba isolates were shown to be responsible for keratitis. CONCLUSION: Genotype T4 was the only Acanthamoeba genotype responsible for keratitis in this study, and represented 79% of non-pathogenic isolates.

Morphogenesis during division and griseofulvin-induced changes of the microtubular cytoskeleton in the parasitic protist, Trichomonas vaginalis.

The behavior of microtubular structures during division was followed by immunofluorescence in Trichomonas vaginalis using an anti-alpha-tubulin monoclonal antibody together with nuclear staining by DAPI, allowing us to describe successive mitotic stages. In contrast to recent reports, we showed that: (1) the microtubular axostyle-pelta complex depolymerized during division, (2) the flagella were assembled during mitosis, and (3) the flagellar number was restored in each daughter kinetid before cytokinesis. Observation of griseofulvin-treated T. vaginalis cells revealed that the elongation of the mitotic spindle or paradesmosis was not the main motile force separating the daughter kinetids to opposite poles during division, suggesting the existence of other mechanisms and/or molecules involved in this morphogenetic event. Examination of treated cells re-incubated in fresh medium showed the nucleation of microtubules radiating from the perinuclear area, the origin of which is discussed. Finally, we confirm the effectiveness of griseofulvin against T. vaginalis and propose that this antifungal drug could be a promising antitrichomonal agent.

Phylogenetic position of the trichomonad parasite of turkeys, Histomonas meleagridis (Smith) Tyzzer, inferred from small subunit rRNA sequence.

The phylogenetic position of the trichomonad, Histomonas meleagridis was determined by analysis of small subunit rRNAs. Molecular trees including all identified parabasalid sequences available in data bases were inferred by distance, parsimony, and likelihood methods. All reveal a close relationship between H. meleagridis, and Dientamoeba fragilis. Moreover, small subunit rRNAs of both amoeboid species have a reduced G + C content and increased chain length relative to other parabasalids. Finally, the rRNA genes from H. meleagridis and D. fragilis share a recent common ancestor with Tritrichomonasfoetus, which exhibits a more developed cytoskeleton. This indicates that Histomonas and Dientamoeba secondarily lost most of the typical trichomonad cytoskeletal structures and hence, do not represent primitive morphologies. A global phylogeny of parabasalids revealed significant discrepancies with morphology-based classifications, such as the polyphyly of most of the parabasalid families and classes included in our study.

Phylogenetic relationships of class II fumarase genes from trichomonad species.

Class II fumarase sequences were obtained by polymerase chain reaction from five trichomonad species. All residues known to be highly conserved in this enzyme were present. Nuclear run-on assays showed that one of the two genes identified in Tritrichomonas foetus was expressed, whereas no fumarase transcripts were detected in the related species Trichomonas vaginalis. These findings corroborate previous biochemical data. Fumarase genes were also expressed in Monocercomonas sp. and Tetratrichomonas gallinarum but not in Pentatrichomonas hominis, Trichomonas gallinae, Trichomonas tenax, and Trichomitus batrachorum under the culture conditions used. Molecular trees inferred by likelihood methods reveal that trichomonad sequences have no affinity to described class II fumarase genes from other eukaryotes. The absence of functional mitochondria in protists such as trichomonads suggests that they diverged from other eukaryotes prior to the alpha-proteobacterial symbiosis that led to mitochondria. Furthermore, they are basal to other eukaryotes in rRNA analyses. However, support for the early-branching status of trichomonads and other amitochondriate protists based on phylogenetic analyses of multiple data sets has been equivocal. Although the presence of hydrogenosomes suggests that trichomonads once had mitochondria, their class II iron-independent fumarase sequences differ markedly from those of other mitochondriate eukaryotes. All of the class II fumarase genes described from other eukaryotes are of apparent alpha-proteobacterial origin and hence a marker of mitochondrial evolution. In contrast, the class II fumarase from trichomonads emerges among other eubacterial homologs. This is intriguing evidence for an independent acquisition of these genes in trichomonads apart from the mitochondrial endosymbiosis event that gave rise to the form present in other eukaryotes. The ancestral trichomonad class II fumarase may represent a prokaryotic form that was replaced in other eukaryotes after the divergence of trichomonads with the movement of endosymbiont genes into the nucleus. Alternatively, it may have been acquired via a separate endosymbiotic event or lateral gene transfer.

Tubulins in Trichomonas vaginalis: molecular characterization of alpha-tubulin genes, posttranslational modifications, and homology modeling of the tubulin dimer.

We have isolated and analysed an alpha-tubulin-encoding gene (atub1) in an early-diverging eukaryote, Trichomonas vaginalis. The complete atub1 open reading frame included 1.356 bp encoding a polypeptide of 452 amino-acyl residues. A second alpha-tubulin gene (atub2) was amplified by PCR using primers derived from consensus alpha-tubulin amino acid sequences. Both T. vaginalis alpha-tubulin sequences showed high identity to those described in other parabasalids (94.4%-97.3%), and exhibited a high degree of similarity to sequences from Metazoa (such as pig brain) and diplomonads (such as Giardia). Despite large evolutionary distances previously observed between trichomonads and mammals, the three-dimensional model of the T. vaginalis tubulin dimer was very similar to that of pig brain. Possible correlations between alpha-tubulin sequences and posttranslational modifications (PTMs) were examined. Our observations corroborated previous data obtained in T. vaginalis using specific anti-PTMs antibodies. As described in the related species Tritrichomonas mobilensis, microtubules are likely acetylated, non-tyrosinated, glutamylated, and non-glycylated in T. vaginalis. Evolutionary considerations concerning the time of appearance of these tubulin PTMs are also discussed since trichomonads are potentially one of the earliest diverging eukaryotic lineages.

Phylogenetic position of parabasalid symbionts from the termite Calotermes flavicollis based on small subunit rRNA sequences.

Small subunit rDNA genes were amplified by polymerase chain reaction using specific primers from mixed-population DNA obtained from the whole hindgut of the termite Calotermes flavicollis. Comparative sequence analysis of the clones revealed two kinds of sequences that were both from parabasalid symbionts. In a molecular tree inferred by distance, parsimony and likelihood methods, and including 27 parabasalid sequences retrieved from the data bases, the sequences of the group II (clones Cf5 and Cf6) were closely related to the Devescovinidae/Calonymphidae species and thus were assigned to the Devescovinidae Foaina. The sequence of the group I (clone Cf1) emerged within the Trichomonadinae and strongly clustered with Tetratrichomonas gallinarum. On the basis of morphological data, the Monocercomonadidae Hexamastix termitis might be the most likely origin of this sequence.

Genetic divergence at the SODA locus of six different formae speciales of Pneumocystis carinii.

Genetic divergence at the SODA (manganese-dependent superoxide dismutase, MnSOD) locus were compared in six Pneumocystis carinii formae speciales isolated from mouse, rabbit, human, macaque and pig. A degenerate oligonucleotide primer strategy was designed to amplify 85-90% of the full-length SODA gene from P. carinii genomic DNA isolates. DNA sequence analysis revealed an A/T bias in the nucleotide composition (71-77.2%) and the presence of seven small introns (41-142 bp), interrupting each P. carinii open reading frame (ORF) at the same position. The MnSOD deduced amino acid sequences from all P. carinii isolates shared residues which were conserved within the MnSOD family and which are required for enzymatic activity and binding of the cofactor metal. Phylogenetic analysis including MnSOD sequences from representatives of the fungal phyla Basidiomycota and Ascomycota indicated that the P. carinii formae speciales form a monophyletic group that is related to the budding yeasts (subphylum Saccharomycotina, previously called class Hemiascomycetes) in the Ascomycota. In the whole Pneumocystis group, P. carinii f. sp. hominis, P. carinii f. sp. macacae and P. carinii f. sp. oryctolagi MnSOD sequences clustered together, as did the rat-derived P. carinii and P. carinii f. sp. muris sequences.

Molecular cloning, expression analysis and iron metal cofactor characterisation of a superoxide dismutase from Toxoplasma gondii.

A genomic region of 12 kb encompassing the gene encoding the superoxide dismutase (SOD) of Toxoplasma gondii has been cloned. The gene contains four exons of 121, 42, 381 and 59 bp which are separated by three introns of 321, 202, and 577 bp, respectively. The open reading frame can be translated into a protein of 201 amino acids with a molecular mass of 22.6 kDa. Alignment indicated that it is a FeSOD, a type only found in bacteria, protozoa and chloroplast of higher plants. Recombinant SOD was expressed in a Escherichia coli double mutant lacking both MnFeSOD and FeSODs. The presence of iron as metal cofactor was confirmed by measurements of iron by absorption mass spectrometry and electron paramagnetic resonance studies. Semi-quantitative reverse transcribed polymerase chain reaction experiments showed a similar amount of SOD transcripts in two developmental stages of T. gondii. Antibodies raised against the purified recombinant protein detected SOD protein in both bradyzoite and tachyzoite forms suggesting this SOD might be essential for the intracellular growth of both developmental stages. Southern blot analysis indicated that SOD occured as a single copy gene in T. gondii genome.

Molecular phylogeny of parabasalids based on small subunit rRNA sequences, with emphasis on the Trichomonadinae subfamily.

We determined small subunit ribosomal DNA sequences from three parabasalid species, Trichomitus batrachorum strain R105, Tetratrichomonas gallinarum, and Pentatrichomonas hominis belonging to the Trichomonadinae subfamily. Unrooted molecular phylogenetic trees inferred by distance, parsimony, and likelihood methods reveal four discrete clades among the parabasalids. The Trichomonadinae form a robust monophyletic group. Within this subfamily T. gallinarum is closely related to Trichomonas species as supported by morphological data, with P. hominis and Pseudotrypanosoma giganteum occupying basal positions. Our analysis does not place T. batrachorum within the Trichomonadinae. Trichomitus batrachorum (strains R105 and BUB) and Hypotrichomonas acosta form a well-separated cluster, suggesting the genus Trichomitus is polyphyletic. The emergence of T. batrachorum precedes the Trichomonadinae-Tritrichomonadinae dichotomy, emphasizing its pivotal evolutionary position among the Trichomonadidae. A third cluster unites the Devescovinidae and the Calonymphidae. The fourth clade contains the three hypermastigid sequences from the genus Trichonympha, which exhibit the earliest emergence among the parabasalids. The addition of these three new parabasalid species did not however resolve ambiguities regarding the relative branching order of the parabasalid clades. The phylogenetic positions of Tritrichomonas faetus, Monocercomonas sp., Dientamoeba fragilis, and the unidentified Reticulitermes flavipes gut symbiont 1 remain unclear.

Analysis of genetic diversity at the iron-containing superoxide dismutase locus in Plasmodium falciparum wild isolates.

In order to investigate the genetic diversity of iron-containing superoxide dismutase (FeSOD) from Plasmodium falciparum, a potential anti-malarial therapeutic target, we cloned and sequenced Plasmodium FeSOD from 26 blood samples from non-infected patients. Fifteen clones had the same nucleotide sequence as that of the FeSOD gene of the P. falciparum strain HB3 cultivated in vitro. The other 11 clones presented mutations responsible for punctual amino acid changes which did not modify key residues for the function or the structure of the enzyme. The high sequence conservation between FeSOD from the isolates confirms that this enzyme could represent a therapeutic target.