S. Fenoy

First Detection and Genotyping of Human-Associated Microsporidia in Pigeons from Urban Parks

ABSTRACT Microsporidia are ubiquitous opportunistic parasites in nature infecting all animal phyla, and the zoonotic potential of this parasitosis is under discussion. Fecal samples from 124 pigeons from seven parks of Murcia (Spain) were analyzed. Thirty-six of them (29.0%) showed structures compatible with microsporidia spores by staining methods. The DNA isolated from 26 fecal samples (20.9%) of microsporidia-positive pigeons was amplified with specific primers for the four most frequent human microsporidia. Twelve pigeons were positive for only Enterocytozoon bieneusi (9.7%), 5 for Encephalitozoon intestinalis (4%), and one for Encephalitozoon hellem (0.8%). Coinfections were detected in eight additional pigeons: E. bieneusi and E. hellem were detected in six animals (4.8%); E. bieneusi was associated with E. intestinalis in one case (0.8%); and E. hellem and E. intestinalis coexisted in one pigeon. No positive samples for Encephalitozoon cuniculi were detected. The internally transcribed spacer genotype could be completed for one E. hellem-positive pigeon; the result was identical to the genotype A1 previously characterized in an E. hellem Spanish strain of human origin. To our knowledge, this is the first time that human-related microsporidia have been identified in urban park pigeons. Moreover, we can conclude that there is no barrier to microsporidia transmission between park pigeons and humans for E. intestinalis and E. hellem. This study is of environmental and sanitary interest, because children and elderly people constitute the main visitors of parks and they are populations at risk for microsporidiosis. It should also contribute to the better design of appropriate prophylactic measures for populations at risk for opportunistic infections.

High-Level Resistance of Nosema ceranae, a Parasite of the Honeybee, to Temperature and Desiccation

ABSTRACT Resistance of Nosema ceranae to different exposure conditions has been evaluated by using Sytox green and DAPI (4′,6-diamidino-2-phenylindole) to test spore viability. High thermotolerance at 60 and 35°C and resistance to desiccation were observed. However, a significant decrease in viability after freezing and a rapid degeneration of spores maintained at 4°C were also detected.

First Cases of Microsporidiosis in Transplant Recipients in Spain and Review of the Literature

ABSTRACT Microsporidia are currently considered emerging pathogens responsible for life-threatening infections in organ transplant recipients. Here, we describe the first cases of intestinal microsporidiosis by Enterocytozoon bieneusi genotype D in two non-HIV-infected renal transplant recipients from Spain. Previously reported cases of microsporidiosis in organ transplant recipients have also been reviewed, highlighting the necessity of considering organ transplant recipients a risk group for microsporidiosis. A systematic search for these parasites is recommended in cases of persistent diarrhea and in the differential diagnosis of other syndromes, such as chronic fever of unknown etiology.

Phylogenetic Approach to the Variability of the Microsporidian Enterocytozoon bieneusi and Its Implications for Inter- and Intrahost Transmission

ABSTRACT Enterocytozoon bieneusi is a microsporidian parasite that infects many vertebrate animals, including humans. The rDNA internal transcribed spacer (ITS) shows a hypervariable sequence; however, so far no clear information has been inferred about strain evolution in this species. We reviewed all the sequences described and performed a phylogenetic study. Four groups of sequences strongly differentiated from each other were detected, although most of the isolates (94%) corresponded to group I. The highly diverse sequences of this group were analyzed using median-joining networks. The host species (humans, pets, swine, cattle, birds, and wild animals) and the continents of origin of the isolates were considered. Central haplotypes in the network were obtained from very diverse hosts and geographical origins. The results show that although E. bieneusi has a broad host specificity, transmission is not completely free: some strains were able to circulate within a given host species and were only occasionally transmitted to another host. Additionally, while not relevant for swine or cattle hosts, geography seems to be a relevant factor for human infection by E. bieneusi.

Detection of zoonotic intestinal parasites in public parks of Spain. Potential epidemiological role of microsporidia.

Several studies have demonstrated that the soil of public parks presents an important source of infection which has a significant impact on public health. Children are the main group affected by accidentally ingestion of contaminated soil. This study was performed in order to identify the presence of zoonotic parasites in dog and cat faecal and soil samples from public parks of Madrid, Spain. Six hundred twenty‐five and seventy‐nine soil and faecal samples (presumably from dogs and cats) respectively were collected from 67 parks. Intestinal parasites were identified in 27 parks (40.3%), which were contamined with Giardia sp. (19.4%), microsporidia (19.4%), Toxocara spp. (16.4%), Cryptosporidium sp. (6%), Entamoeba histolytica (3%) and Ancylostomidae (3%). Combinations of two or more intestinal parasites were found in 11 parks, and it was common to find Giardia and microsporidia together in samples. Intestinal parasites were detected in 18% (112/625) of soil samples. The most frequent parasite species found in the examined soil samples were Toxocara spp. (16.4%), followed by Giardia sp. (4.5%) and Strongyloides sp. larvae (3%). The zoonotic parasites found in the 79 faecal samples were Giardia sp. (17.7%), Cryptosporidium sp. (9%), E. histolytica (2.5%), Trichuris vulpis (1.3%), Toxascaris leonina (1.3%) and microsporidia spores (28%). Microsporidia characterization by amplification of DNA confirmed 10 samples as positive, eight for E. bieneusi and two for E. hellem by PCR. The role of those parasites in the environment are discussed.

Intraspecies Genotype Variability of the Microsporidian Parasite Encephalitozoon hellem

ABSTRACT Seven isolates of Encephalitozoon hellem from human immunodeficiency virus-positive patients were genotyped through a series of markers: the internal transcribed spacer (ITS) of ribosomal DNA, the polar tube protein (PTP) gene, and two intergenic spacers (IGS-TH and IGS-HZ) whose polymorphism is newly reported. The genome markers were all analyzed at three levels: PCR amplification followed by polyacrylamide gel electrophoresis, single-strand conformation analysis (SSCA), and DNA sequencing. The polymorphisms detected involve insertions/deletions and point mutations. SSCA can distinguish any pair of sequences, even those differing by a single base pair. The different isolates studied fit into the previously described ITS genotype 1A, except one which seems to be a 2A derivative variant (2D). When PTP and the new markers IGS-TH and IGS-HZ were analyzed, most of the isolates displayed different genotypes, demonstrating that E. hellem has a strong intraspecies variability. A set of markers such as those used here may be very useful in genotyping of clinical samples and in the assessment of epidemiological relationships among E. hellem strains.

Detection of Enterocytozoon bieneusi in Two Human Immunodeficiency Virus—Negative Patients with Chronic Diarrhea by Polymerase Chain Reaction in Duodenal Biopsy Specimens and Review

Intestinal microsporidiosis has been associated traditionally with severely immunocompromised patients with AIDS. We describe two new cases of intestinal microsporidiosis due to Enterocytozoon bieneusi in human immunodeficiency virus-negative adults. Both patients presented with chronic nonbloody diarrhea, and one had intestinal lymphangiectasia as well. Intestinal microsporidiosis was diagnosed by evaluation of stool samples, and the specific species was determined by use of polymerase chain reaction (PCR) in duodenal biopsy specimens. To our knowledge, this is the first report of confirmation of E. bieneusi in the intestinal epithelium of HIV-negative individuals by use of PCR in duodenal biopsy specimens. Cases of intestinal microsporidiosis in HIV-negative individuals reported in the English-language literature are reviewed. These two new cases along with those described previously corroborate the need to evaluate for microsporidia in HIV-negative individuals with unexplained diarrhea.

Molecular Characterization of Human-Pathogenic Microsporidia and Cyclospora cayetanensis Isolated from Various Water Sources in Spain: a Year-Long Longitudinal Study

ABSTRACT Recent studies suggest the involvement of water in the epidemiology of Cyclospora cayetanensis and some microsporidia. A total of 223 samples from four drinking water treatment plants (DWTPs), seven wastewater treatment plants (WWTPs), and six locations of influence (LI) on four river basins from Madrid, Spain, were analyzed from spring 2008 to winter 2009. Microsporidia were detected in 49% of samples (109/223), Cyclospora spp. were detected in 9% (20/223), and both parasites were found in 5.4% (12/223) of samples. Human-pathogenic microsporidia were detected, including Enterocytozoon bieneusi (C, D, and D-like genotypes), Encephalitozoon intestinalis, Encephalitozoon cuniculi (genotypes I and III), and Anncaliia algerae. C. cayetanensis was identified in 17 of 20 samples. To our knowledge, this is the first study that shows a year-long longitudinal study of C. cayetanensis in drinking water treatment plants. Additionally, data about the presence and molecular characterization of the human-pathogenic microsporidia in drinking water, wastewater, and locations of influence during 1 year in Spain are shown. It is noteworthy that although the DWTPs and WWTPs studied meet European and national regulations on water sanitary quality, both parasites were found in water samples from these plants, supporting the idea that new and appropriate controls and regulations for drinking water, wastewater, and recreational waters should be proposed to avoid health risks from these pathogens.

In Vitro Culture, Ultrastructure, Antigenic, and Molecular Characterization of Encephalitozoon cuniculi Isolated from Urine and Sputum Samples from a Spanish Patient with AIDS

ABSTRACT In this report we describe the cultivation of two isolates of microsporidia, one from urine and the other from sputum samples from a Spanish AIDS patient. We identified them as Encephalitozoon cuniculi, type strain III (the dog genotype), based on ultrastructure, antigenic characteristics, PCR, and the sequence of the ribosomal DNA internal transcribed spacer region.

Novel Acanthamoeba 18S rRNA gene sequence type from an environmental isolate

The free-living amoebae, Acanthamoeba, can act as opportunistic parasites on a wide range of vertebrates and are becoming a serious threat to human health due to the resistance of their cysts to harsh environmental conditions, disinfectants, some water treatment practices, and their ubiquitous distribution. Subgenus classification based on morphology is being replaced by a classification based on the sequences of the 18S rRNA gene with a total of 18 different genotypes (T1–T18). A new environmental strain of Acanthamoeba isolated from a waste water treatment plant is presented in this study as a candidate for the description of the novel genotype T19 after phylogenetic analysis.