-
MethodsX Dec 2023A wide range of intestinal protozoan parasites inhabit the human gut. To establish a more comprehensive molecular screening, we designed PCR-sequencing screening methods...
A wide range of intestinal protozoan parasites inhabit the human gut. To establish a more comprehensive molecular screening, we designed PCR-sequencing screening methods for spp., including commensal species, and , and performed such methods using 174 stool samples collected from Kenyan children. The prevalences of the target species were as follows: (2/174, 1.1%), (20/174, 11.5%), (107/174, 61.5%), (77/174, 44.3%), and (54/174, 31.0%). PCR amplicons specific to was differentiated to assemblages A (8/174, 4.6%) and B (46/174, 26.4%). PCR specificity for spp. was quite high, except for some cross-reactions between detection primers and although the false-positive amplicons were discernible by the band size. The 18S rRNA PCR primers that was designed by Monis et al. in 1999 for , have specificity issue, therefore amplicon sequencing was essential not only to determine assemblage classifications but also to confirm the positive results by eliminating potential non-specific reactions. The detection sensitivity of both the universal PCR and the PCR was more than 100 copies of the target loci, which is sufficient for detecting a single trophozoite or cyst of both species.
PubMed: 37744888
DOI: 10.1016/j.mex.2023.102361 -
Memorias Do Instituto Oswaldo Cruz Feb 2016This study aimed to estimate the frequency, associated factors, and molecular characterisation of Entamoeba histolytica, Entamoeba dispar, Entamoeba moshkovskii,...
Frequency and molecular characterisation of Entamoeba histolytica, Entamoeba dispar, Entamoeba moshkovskii, and Entamoeba hartmanni in the context of water scarcity in northeastern Brazil.
This study aimed to estimate the frequency, associated factors, and molecular characterisation of Entamoeba histolytica, Entamoeba dispar, Entamoeba moshkovskii, andEntamoeba hartmanni infections. We performed a survey (n = 213 subjects) to obtain parasitological, sanitation, and sociodemographic data. Faecal samples were processed through flotation and centrifugation methods.E. histolytica, E. dispar, E. moshkovskii, and E. hartmanni were identified by nested-polymerase chain reaction (PCR). The overall prevalence of infection was 22/213 (10.3%). The infection rate among subjects who drink rainwater collected from roofs in tanks was higher than the rate in subjects who drink desalinated water pumped from wells; similarly, the infection rate among subjects who practice open defecation was significantly higher than that of subjects with latrines. Out of the 22 samples positive for morphologically indistinguishableEntamoeba species, the differentiation by PCR was successful for 21. The species distribution was as follows: 57.1% to E. dispar, 23.8% to E. histolytica, 14.3% toE. histolytica and E. dispar, and 4.8% E. dispar and E. hartmanni. These data suggest a high prevalence of asymptomatic infection by the group of morphologically indistinguishable Entamoeba histolytica/dispar/moshkovskiicomplex and E. hartmanni species. In this context of water scarcity, the sanitary and socioenvironmental characteristics of the region appear to favour transmission.
Topics: Adolescent; Adult; Aged; Aged, 80 and over; Brazil; Child; Cross-Sectional Studies; DNA, Protozoan; Drinking Water; Droughts; Entamoeba; Entamoebiasis; Feces; Female; Humans; Male; Middle Aged; Molecular Typing; Polymerase Chain Reaction; Poverty; Prevalence; Water Wells; Young Adult
PubMed: 26841049
DOI: 10.1590/0074-02760150383 -
Parasites & Vectors Mar 2021Entamoeba species harbored by humans have different degrees of pathogenicity. The present study explores the intra- and interspecific diversity, phylogenetic...
BACKGROUND
Entamoeba species harbored by humans have different degrees of pathogenicity. The present study explores the intra- and interspecific diversity, phylogenetic relationships, prevalence and distribution of tetra- and octonucleated cyst-producing Entamoeba in different Brazilian regions.
METHODS
Cross-sectional studies were performed to collect fecal samples (n = 1728) and sociodemographic data in communities located in four Brazilian biomes: Atlantic Forest, Caatinga, Cerrado, and Amazon. Fecal samples were subjected to molecular analysis by partial small subunit ribosomal DNA sequencing (SSU rDNA) and phylogenetic analysis.
RESULTS
Light microscopy analysis revealed that tetranucleated cysts were found in all the studied biomes. The highest positivity rates were observed in the age group 6-10 years (23.21%). For octonucleated cysts, positivity rates ranged from 1 to 55.1%. Sixty SSU rDNA Entamoeba sequences were obtained, and four different species were identified: the octonucleated E. coli, and the tetranucleated E. histolytica, E. dispar, and E. hartmanni. Novel haplotypes (n = 32) were characterized; however, new ribosomal lineages were not identified. The Entamoeba coli ST1 subtype predominated in Atlantic Forest and Caatinga, and the ST2 subtype was predominant in the Amazon biome. E. histolytica was detected only in the Amazon biome. In phylogenetic trees, sequences were grouped in two groups, the first containing uni- and tetranucleated and the second containing uni- and octonucleated cyst-producing Entamoeba species. Molecular diversity indexes revealed a high interspecific diversity for tetra- and octonucleated Entamoeba spp. (H ± SD = 0.9625 ± 0.0126). The intraspecific diversity varied according to species or subtype: E. dispar and E. histolytica showed lower diversity than E. coli subtypes ST1 and ST2 and E. hartmanni.
CONCLUSIONS
Tetra- and octonucleated cyst-producing Entamoeba are endemic in the studied communities; E. histolytica was found in a low proportion and only in the Amazon biome. With regard to E. coli, subtype ST2 was predominant in the Amazon biome. The molecular epidemiology of Entamoeba spp. is a field to be further explored and provides information with important implications for public health.
Topics: Adolescent; Brazil; Child; Child, Preschool; Cross-Sectional Studies; DNA, Protozoan; Ecosystem; Entamoeba; Entamoebiasis; Feces; Female; Genetic Variation; Humans; Infant; Infant, Newborn; Male; Phylogeny; Prevalence; Sequence Analysis, DNA
PubMed: 33731176
DOI: 10.1186/s13071-021-04672-y -
Clinical Microbiology Reviews Jul 2007The genus Entamoeba contains many species, six of which (Entamoeba histolytica, Entamoeba dispar, Entamoeba moshkovskii, Entamoeba polecki, Entamoeba coli, and Entamoeba... (Review)
Review
The genus Entamoeba contains many species, six of which (Entamoeba histolytica, Entamoeba dispar, Entamoeba moshkovskii, Entamoeba polecki, Entamoeba coli, and Entamoeba hartmanni) reside in the human intestinal lumen. Entamoeba histolytica is the causative agent of amebiasis and is considered a leading parasitic cause of death worldwide in humans. Although recent studies highlight the recovery of E. dispar and E. moshkovskii from patients with gastrointestinal symptoms, there is still no convincing evidence of a causal link between the presence of these two species and the symptoms of the host. New approaches to the identification of E. histolytica are based on detection of E. histolytica-specific antigen and DNA in stool and other clinical samples. Several molecular diagnostic tests, including conventional and real-time PCR, have been developed for the detection and differentiation of E. histolytica, E. dispar, and E. moshkovskii in clinical samples. The purpose of this review is to discuss different methods that exist for the identification of E. histolytica, E. dispar, and E. moshkovskii which are available to the clinical diagnostic laboratory. To address the need for a specific diagnostic test for amebiasis, a substantial amount of work has been carried out over the last decade in different parts of the world. The molecular diagnostic tests are increasingly being used for both clinical and research purposes. In order to minimize undue treatment of individuals infected with other species of Entamoeba such as E. dispar and E. moshkovskii, efforts have been made for specific diagnosis of E. histolytica infection and not to treat based simply on the microscopic examination of Entamoeba species in the stool. The incorporation of many new technologies into the diagnostic laboratory will lead to a better understanding of the public health problem and measures to control the disease.
Topics: Animals; Antibodies, Protozoan; Antigens, Protozoan; Chromatography; DNA, Protozoan; Entamoeba; Entamoebiasis; Enzyme-Linked Immunosorbent Assay; Humans; Isoenzymes; Nucleic Acid Amplification Techniques; Oligonucleotide Array Sequence Analysis; Polymerase Chain Reaction; Protozoan Proteins
PubMed: 17630338
DOI: 10.1128/CMR.00004-07 -
Food and Waterborne Parasitology Jun 2021is the only pathogenic species of the genus and is morphologically identical to / ( complex) hence cannot be microscopically differentiated. The other spp. found in...
is the only pathogenic species of the genus and is morphologically identical to / ( complex) hence cannot be microscopically differentiated. The other spp. found in humans ( and ) can be differentiated morphologically from this complex. However, some of their morphologic features overlap making differential diagnosis difficult. This study aimed at determining the occurrence of spp. in patients seeking treatment for diarrhea and/or abdominal discomfort at two clinics in Mukuru informal settlement in Nairobi, Kenya. Faecal samples were collected from 895 patients, examined microscopically following direct wet smear and formal-ether concentration methods. spp. positive faecal samples were subjected to DNA extraction and species-specific nested polymerase chain reaction of the 18S ribosomal RNA (rRNA). By microscopy, spp. cysts or trophozoites were detected in 114/895 (12.7%, 95% Confidence Interval (CI) 10.6-15.1) faecal samples. By nested PCR, the prevalence was: (7.5%, 95% CI 5.9-9.4, 67/895) and (8.2%, 95% CI 6.5-10.2, 73/895). Among the spp. complex positive samples, nested PCR detected and DNA in 63/114 (55.3%) and 37/114 (32.5%), samples respectively. Among the / PCR negative samples (32.5%), 21 (18.4%) contained cysts of either (19) or (2) by nested PCR. spp. infections were most common among participants aged 21-30 years; however it was not significant ( = 0.7). spp. infections showed an inverse relationship with diarrhea being most common among participants without diarrhea ( = 0.0). The difference was significant for ( = 0.0) but not significant for ( = 0.1). Only infections were significantly associated with sex ( = 0.0). This study highlights the need for differentiation of from other spp. by molecular tools for better management of amoebiasis.
PubMed: 33898796
DOI: 10.1016/j.fawpar.2021.e00122 -
Parasite Epidemiology and Control May 2020Application of next-generation sequencing (NGS) to genomic DNA extracted from sewage offers a unique and cost-effective opportunity to study the genetic diversity of...
BACKGROUND
Application of next-generation sequencing (NGS) to genomic DNA extracted from sewage offers a unique and cost-effective opportunity to study the genetic diversity of intestinal parasites. In this study, we used amplicon-based NGS to reveal and differentiate several common luminal intestinal parasitic protists, specifically , , , and , in sewage samples from Swedish treatment plants.
MATERIALS AND METHODS
Influent sewage samples were subject to gradient centrifugation, DNA extraction and PCR-based amplification using three primer pairs designed for amplification of eukaryotic nuclear 18S ribosomal DNA. PCR products were sequenced using ILLUMINA® technology, and resulting sequences were annotated to species and subtype level using the in-house BION software, sequence clustering, and phylogenetic analysis.
RESULTS
A total of 26 samples from eight treatment plants in central/southern Sweden were analysed. sp. and were detected in all samples, and most samples ( = 20) were positive for . Moreover, we detected , , , , and in 1, 11, 4, 10, and 7 samples, respectively. The level of genetic divergence observed within and was 20.2% and 7.7%, respectively, across the ~400-bp region studied, and two clades of were found. As expected, sp. subtypes 1-4 were present in almost all samples; however, ST8 was present in 10 samples and was the only subtype not commonly found in humans that was present in multiple samples.
CONCLUSIONS
and were identified as universal members of the "sewage microbiome". sp. ST8, which has been rarely reported in humans, was a very common finding, indicating that a hitherto unidentified but common host of ST8 contributed to the sewage influent. The study also provided substantial new insight into the intra-generic diversity of and .
PubMed: 31909230
DOI: 10.1016/j.parepi.2019.e00131 -
Parasite Epidemiology and Control May 2022Comprehensive detection and differentiation of intestinal protists mostly rely on DNA-based methods. Here, we evaluated next-generation sequencing of eukaryotic nuclear...
Comprehensive detection and differentiation of intestinal protists mostly rely on DNA-based methods. Here, we evaluated next-generation sequencing of eukaryotic nuclear ribosomal genes (metabarcoding) for the detection and differentiation of intestinal eukaryotic protists in the stool of healthy Tunisian individuals. Thirty-six faecal DNA samples previously evaluated by microscopy and ameboid species-specific PCRs were tested. The hypervariable regions V3-V4 and V3-V5 of the 18S rRNA gene were amplified using three universal eukaryotic primer sets and sequenced using Illumina®MiSeq sequencing. In addition, real-time PCR assays were used to detect , , and spp. The metabarcoding assay detected (subtypes 1, 2, and 3) and archamoebid species and subtypes (, , RL1 and RL2, , RL1) in 27 (75%) and 22 (61%) of the 36 stool samples, respectively. Meanwhile, the assay had limited sensitivity for flagellates as evidenced by the fact that no -specific reads were found in any of the five -positive samples included, and -specific reads were observed only in 3/13 -positive samples. None of the samples were positive for by any of the methods. In conclusion, a large variety of intestinal eukaryotic protists were detected and differentiated at species and subtype level; however, limited sensitivity for common flagellates was observed.
PubMed: 35146142
DOI: 10.1016/j.parepi.2022.e00242 -
MSphere Aug 2021An estimated 3.5 billion people are colonized by intestinal parasites worldwide. Intestinal parasitic eukaryotes interact not only with the host but also with the...
Exposure to Parasitic Protists and Helminths Changes the Intestinal Community Structure of Bacterial Communities in a Cohort of Mother-Child Binomials from a Semirural Setting in Mexico.
An estimated 3.5 billion people are colonized by intestinal parasites worldwide. Intestinal parasitic eukaryotes interact not only with the host but also with the intestinal microbiota. In this work, we studied the relationship between the presence of multiple enteric parasites and the community structures of gut bacteria and eukaryotes in an asymptomatic mother-child cohort from a semirural community in Mexico. Fecal samples were collected from 46 mothers and their respective children, with ages ranging from 2 to 20 months. Mothers and infants were found to be multiparasitized by Blastocystis hominis, Entamoeba dispar, Endolimax nana, Chilomastix mesnili, Iodamoeba butshlii, Entamoeba coli, Hymenolepis nana, and Ascaris lumbricoides. Sequencing of bacterial 16S rRNA and eukaryotic 18S rRNA genes showed a significant effect of parasite exposure on bacterial beta-diversity, which explained between 5.2% and 15.0% of the variation of the bacterial community structure in the cohort. Additionally, exposure to parasites was associated with significant changes in the relative abundances of multiple bacterial taxa, characterized by an increase in and decreases in and . Parasite exposure was not associated with changes in intestinal eukaryote relative abundances. However, we found several significant positive correlations between intestinal bacteria and eukaryotes, including with and Prevotella stercorea with Entamoeba hartmanni, as well as the co-occurrence of the fungus with and , , and Prevotella copri and the fungus with . The parasitic exposure-associated changes in the bacterial community structure suggest effects on microbial metabolic routes, host nutrient uptake abilities, and intestinal immunity regulation in host-parasite interactions. The impact of intestinal eukaryotes on the prokaryotic microbiome composition of asymptomatic carriers has not been extensively explored, especially in infants and mothers with multiple parasitic infections. In this work, we studied the relationship between protist and helminth parasite colonization and the intestinal microbiota structure in an asymptomatic population of mother-child binomials from a semirural community in Mexico. We found that the presence of parasitic eukaryotes correlated with changes in the bacterial gut community structure in the intestinal microbiota in an age-dependent way. Parasitic infection was associated with an increase in the relative abundance of the class and decreases of and . Parasitic infection was not associated with changes in the eukaryote community structure. However, we observed strong positive correlations between bacterial and other eukaryote taxa, identifying novel relationships between prokaryotes and fungi reflecting interkingdom interactions within the human intestine.
Topics: Adolescent; Adult; Animals; Bacteria; Cohort Studies; Feces; Female; Gastrointestinal Microbiome; Helminths; Host-Parasite Interactions; Humans; Infant; Intestinal Diseases, Parasitic; Mexico; Middle Aged; Models, Statistical; Mothers; Parasites; RNA, Ribosomal, 16S; Rural Population; Young Adult
PubMed: 34406855
DOI: 10.1128/mSphere.00083-21 -
Brazilian Journal of Biology = Revista... Aug 2018Intestinal parasitic infections in immunocompromised patients can lead to serious complications when not diagnosed and treated early. This study aimed to investigate the...
Intestinal parasitic infections in immunocompromised patients can lead to serious complications when not diagnosed and treated early. This study aimed to investigate the frequency of intestinal parasites in cancer patients undergoing chemotherapy in the South of Brazil. Three fecal samples collected from each patient (73 individuals) were processed by Ritchie and Faust techniques and submitted to specific staining methods for intestinal protozoa. A 61.6% parasite and/or commensal positivity was found. Helminths identified were Ascaris lumbricoides (33.3%), Taenia spp. (6.6%), Strongyloides stercoralis (4.4%) and Trichuris trichiura (2.2%). Among protozoans, Giardia lamblia (26.6%), Cryptosporidium spp. (13.3%) and Cystoisospora belli (4.4%) were identified. The presence of Entamoeba coli, Endolimax nana and Entamoeba hartmanni was also recorded. The results obtained warn of the importance of fecal parasitological diagnosis and the use of specific staining methods for the detection of intestinal parasites in cancer patients. These exams should be regularly requested at the patient's first clinic visit, given the high prevalence found in this study and the possible severity of such conditions for these individuals.
Topics: Adult; Aged; Aged, 80 and over; Animals; Brazil; Comorbidity; Cryptosporidium; Entamoeba; Feces; Female; Giardia lamblia; Helminths; Humans; Intestinal Diseases, Parasitic; Male; Middle Aged; Neoplasms; Prevalence; Young Adult
PubMed: 29185612
DOI: 10.1590/1519-6984.175364 -
The American Journal of Tropical... Jan 2021Giardia intestinalis is one of the most common causes of parasite-induced diarrhea, abdominal pain, flatulence, and malabsorption. Yet, data on the epidemiology of G.... (Comparative Study)
Comparative Study
Giardia intestinalis is one of the most common causes of parasite-induced diarrhea, abdominal pain, flatulence, and malabsorption. Yet, data on the epidemiology of G. intestinalis infections in North Africa are limited. The purpose of this study was to carry out a retrospective survey on the level of intestinal parasitism with a particular emphasis on G. intestinalis in children and adults in Algiers, Algeria. A total of 2,054 individuals from outpatient clinics or hospitalized at Beni-Messous University Hospital of Algiers undergoing stool microscopy for ova and parasites were included. The overall parasite infection rate was 28%. In the 567 parasite-positive samples, Blastocystis was found most frequently (57.3%), followed in frequency by Endolimax nana (41.0%), Entamoeba histolytica/dispar (19.6%), G. intestinalis (17.1%), Entamoeba coli (13.9%), Chilomastix mesnili (1.0%), Iodamoeba bütschlii (0.7%), Entamoeba hartmanni (0.5%), and Cryptosporidium spp. (0.2%). Intestinal parasites were generally more common in adults than in children, except for Giardia, which was more common in children (P = 0.0001). Giardia infection was independent of gender (P = 0.94). Compared with other intestinal parasitic infections, clinical manifestations, such as abdominal pain (P = 0.28) and diarrhea (P = 0.82), were found not to be significantly linked to Giardia infection. In conclusion, G. intestinalis is common in individuals referred to the University Hospital of Beni-Messous with digestive symptoms, particularly so in children. However, in our study, intestinal symptoms appeared not to be more linked to Giardia than to other intestinal parasites.
Topics: Adolescent; Adult; Age Factors; Aged; Aged, 80 and over; Algeria; Animals; Child; Child, Preschool; Epidemiological Monitoring; Feces; Female; Giardia; Giardiasis; Humans; Infant; Infant, Newborn; Intestinal Diseases, Parasitic; Male; Middle Aged; Prevalence; Retrospective Studies; Sex Factors; Young Adult
PubMed: 33534771
DOI: 10.4269/ajtmh.20-0187