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1.
Am J Trop Med Hyg ; 110(3): 460-469, 2024 Mar 06.
Article in English | MEDLINE | ID: mdl-38266286

ABSTRACT

Zoonotic human infections with Ancylostoma ceylanicum have recently been reported in the Americas. We used archived human stool samples to study the geographic distribution of human infections with A. ceylanicum and anthropophilic hookworms in different geoclimatic regions (coastal, Andean, and Amazon) of Ecuador. We analyzed retrospectively archived human stool samples from five studies previously screened for hookworm infection by microscopy, of which four included hookworm-positive samples only and one involved hookworm-negative samples to increase geographic distribution of sampling. Stools were analyzed using multi-parallel quantitative polymerase chain reaction (qPCR) assays to detect Necator americanus, Ancylostoma duodenale, A. ceylanicum, Ascaris lumbricoides, Trichuris trichiura, and Strongyloides stercoralis. Sequencing was done for the A. ceylanicum cox1 gene. A total of 132 samples were analyzed, of which 69 (52.3%) were from hookworm-positive and 63 (47.7%) from hookworm-negative individuals by microscopy. Overall, 82.6% of microscopy-positive samples and 33.3% of microscopy-negative samples were positive for hookworm by qPCR. Of microscopy-positive samples, 36.2% were A. ceylanicum, 37.7% A. duodenale, and 33.3% N. americanus, whereas equivalent proportions for microscopy-negative samples were 1.6%, 31.7%, and 1.6%, respectively. Ancylostoma duodenale was the most widely dispersed geographically, followed by N. americanus. Ancylostoma ceylanicum was least dispersed but was detected in coastal and Amazon regions. In conclusion, human infections with A. ceylanicum, A. duodenale, and N. americanus were detected in different geoclimatic regions of Ecuador. Additional studies are required to further define the epidemiology of human A. ceylanicum infections, but the potentially widespread presence of this helminth in human populations in Ecuador has implications for hookworm control strategies.


Subject(s)
Ancylostomiasis , Hookworm Infections , Animals , Humans , Ancylostoma/genetics , Ancylostomatoidea , Ancylostomiasis/epidemiology , Ancylostomiasis/diagnosis , Retrospective Studies , Ecuador/epidemiology , Hookworm Infections/epidemiology , Zoonoses/epidemiology , Feces
2.
PLoS Negl Trop Dis ; 17(11): e0011777, 2023 Nov.
Article in English | MEDLINE | ID: mdl-37983257

ABSTRACT

BACKGROUND: There are limited longitudinal data on the acquisition of Giardia lamblia infections in childhood using molecular assays to detect and type assemblages, and measure effects of infections on diarrhea risk and childhood growth. METHODS: We analysed stool samples from a surveillance sample within a birth cohort in a rural district in tropical Ecuador. The cohort was followed to 8 years of age for the presence of G. lamblia in stools by quantitative PCR and A and B assemblages by Taqman assay or Sanger sequencing. We explored risk factors associated with infection using generalized estimating equations applied to longitudinal binary outcomes, and longitudinal panel data analysis to estimate effects of infection on diarrhea and growth trajectories. RESULTS: 2,812 stool samples collected between 1 month and 8 years of age from 498 children were analyzed and showed high rates of infection: 79.7% were infected at least once with peak prevalence (53.9%) at 5 years. Assemblage B was accounted for 56.8% of genotyped infections. Risk factors for infection included male sex (P = 0.001), daycare attendance (P<0.001), having a household latrine (P = 0.04), childhood (P<0.001) and maternal soil-transmitted helminth (P = 0.029) infections, and exposures to donkeys (age interaction P = 0.034). G. lamblia was associated with increased risk of diarrhea (per episode, RR 1.03, 95% CI 1.01-1.06, P = 0.011) during the first 3 years of life and a transient impairment of weight (age interaction P = 0.017) and height-for-age (age interaction P = 0.025) trajectories between 1 and 4 years of age. There was no increased risk of either assemblage being associated with outcomes. CONCLUSION: Our data show a relatively high edemicity of G. lamblia transmission during childhood in coastal Ecuador, and evidence that infection is associated with a transiently increased risk of diarrhea during the first 3 years of life and impairment of weight and height between 1 and 4 years.


Subject(s)
Giardia lamblia , Giardiasis , Child , Humans , Male , Child, Preschool , Infant, Newborn , Giardiasis/epidemiology , Giardia lamblia/genetics , Birth Cohort , Ecuador/epidemiology , Giardia/genetics , Diarrhea/epidemiology , Feces
3.
Parasit Vectors ; 13(1): 141, 2020 Mar 18.
Article in English | MEDLINE | ID: mdl-32188497

ABSTRACT

BACKGROUND: Enteric parasites are transmitted in households but few studies have sampled inside households for parasites and none have used sensitive molecular methods. METHODS: We collected bed and living room dust samples from households of children participating in a clinical trial of anthelmintic treatment in rural coastal Ecuador. Dust was examined for presence of DNA specific for 11 enteric parasites (Ascaris lumbricoides, Trichuris trichiura, Ancylostoma duodenale, Necator americanus, Strongyloides stercoralis, Toxocara canis and T. cati, Giardia lamblia, Blastocystis hominis, Cryptosporidium spp., and Entamoeba histolytica) by quantitative PCR (qPCR). RESULTS: Of the 38 households sampled, 37 had positive dust for at least one parasite and up to 8 parasites were detected in single samples. Positivity was greatest for B. hominis (79% of household samples) indicating a high level of environmental fecal contamination. Dust positivity rates for individual pathogens were: S. stercoralis (52%), A. lumbricoides (39%), G. lamblia (39%), Toxocara spp. (42%), hookworm (18%) and T. trichiura (8%). DNA for Cryptosporidium spp. and E. histolytica was not detected. Bed dust was more frequently positive than floor samples for all parasites detected. Positivity for A. lumbricoides DNA in bed (adjusted OR: 10.0, 95% CI: 2.0-50.1) but not floor dust (adjusted OR: 3.6, 95% CI: 0.3-37.9) was significantly associated with active infections in children. CONCLUSIONS: To our knowledge, this is the first use of qPCR on environmental samples to detect a wide range of enteric pathogen DNA. Our results indicate widespread contamination of households with parasite DNA and raise the possibility that beds, under conditions of overcrowding in a humid tropical setting, may be a source of transmission.


Subject(s)
Beds/parasitology , DNA/analysis , Dust/analysis , Family Characteristics , Intestinal Diseases, Parasitic/transmission , Parasites/genetics , Adolescent , Animals , Child , Controlled Clinical Trials as Topic , Environment , Feces/parasitology , Female , Humans , Male , Real-Time Polymerase Chain Reaction , Rural Population
4.
Methods Mol Biol ; 1745: 277-286, 2018.
Article in English | MEDLINE | ID: mdl-29476474

ABSTRACT

Parasites counteract the action of the immune system and other environmental pressures by modulating and changing the composition of their cell surfaces. Surface multigene protein families are defined not only by highly variable regions in length and/or sequence exposed to the outer space but also by conserved sequences codifying for the signal peptide, hydrophobic C-terminal regions necessary for GPI modifications, as well as conserved UTR regions for mRNA regulation. The method here presented exploits these conserved signatures for characterizing variations in the mRNA expression of clonal cell populations of protozoan parasites using a combination of nested PCR amplification and capillary electrophoresis. With this workflow, in silico gels from isolated cell clones can be generated, thus providing an excellent tool for analyzing cellular heterogeneity in protozoan parasites.


Subject(s)
DNA Fingerprinting , Genes, Protozoan , Membrane Proteins/genetics , Multigene Family , Parasites/genetics , Polymerase Chain Reaction , Animals , Conserved Sequence , DNA Fingerprinting/methods , Databases, Nucleic Acid , Electrophoresis, Capillary , Polymerase Chain Reaction/methods
5.
Open Biol ; 5(12): 150190, 2015 Dec.
Article in English | MEDLINE | ID: mdl-26674416

ABSTRACT

Trypanosoma cruzi has a complex life cycle comprising pools of cell populations which circulate among humans, vectors, sylvatic reservoirs and domestic animals. Recent experimental evidence has demonstrated the importance of clonal variations for parasite population dynamics, survival and evolution. By limiting dilution assays, we have isolated seven isogenic clonal cell lines derived from the Pan4 strain of T. cruzi. Applying different molecular techniques, we have been able to provide a comprehensive characterization of the expression heterogeneity in the mucin-associated surface protein (MASP) gene family, where all the clonal isogenic populations were transcriptionally different. Hierarchical cluster analysis and sequence comparison among different MASP cDNA libraries showed that, despite the great variability in MASP expression, some members of the transcriptome (including MASP pseudogenes) are conserved, not only in the life-cycle stages but also among different strains of T. cruzi. Finally, other important aspects for the parasite, such as growth, spontaneous metacyclogenesis or excretion of different catabolites, were also compared among the clones, demonstrating that T. cruzi populations of cells are also phenotypically heterogeneous. Although the evolutionary strategy that sustains the MASP expression polymorphism remains unknown, we suggest that MASP clonal variability and phenotypic heterogeneities found in this study might provide an advantage, allowing a rapid response to environmental pressure or changes during the life cycle of T. cruzi.


Subject(s)
Membrane Proteins/metabolism , Protozoan Proteins/metabolism , Trypanosoma cruzi/metabolism , Membrane Proteins/genetics , Protozoan Proteins/genetics , Trypanosoma cruzi/genetics
6.
Trends Parasitol ; 31(8): 350-6, 2015 Aug.
Article in English | MEDLINE | ID: mdl-26070403

ABSTRACT

Some of the most crucial phenotypic aspects of parasites, such as an antigen-coated surface, parasite sexual differentiation, virulence, and drug resistance, rely on adaptive plasticity and/or stochastic events. At a population level, cell to cell variability represents an avenue for rapid response to drastic changes in the environment. Single cell approaches can be used to unravel the different strategies used by parasites to survive in the context of regulated transcriptional control (apicomplexa) or in its absence (kinetoplastids).


Subject(s)
Host-Parasite Interactions/physiology , Protozoan Infections/parasitology , Animals , Biological Evolution , Host-Parasite Interactions/genetics , Humans , Parasitology/trends , Phenotype , Single-Cell Analysis
7.
Infect Immun ; 79(10): 3993-4001, 2011 Oct.
Article in English | MEDLINE | ID: mdl-21788387

ABSTRACT

We describe the characterization, purification, expression, and location of a 52-kDa protein secreted during interaction between the metacyclic form of Trypanosoma cruzi and its target host cell. The protein, which we have named MASP52, belongs to the family of mucin-associated surface proteins (MASPs). The highest levels of expression of both the protein and mRNA occur during the metacyclic and bloodstream trypomastigote stages, the forms that infect the vertebrate host cells. The protein is located in the plasma membrane and in the flagellar pockets of the epimastigote, metacyclic, and trypomastigote forms and is secreted into the medium at the point of contact between the parasite and the cell membrane, as well as into the host-cell cytosol during the amastigote stage. IgG antibodies specific against a synthetic peptide corresponding to the catalytic zone of MASP52 significantly reduce the parasite's capacity to infect the host cells. Furthermore, when the protein is adsorbed onto inert particles of bentonite and incubated with a nonphagocytic cell culture, the particles are able to induce endocytosis in the cells, which seems to demonstrate that MASP52 plays a role in a process whereby the trypomastigote forms of the parasite invade the host cell.


Subject(s)
Gene Expression Regulation, Developmental , Life Cycle Stages , Membrane Proteins/metabolism , Protozoan Proteins/metabolism , Trypanosoma cruzi/growth & development , Amino Acid Sequence , Animals , Cell Membrane/parasitology , Chlorocebus aethiops , Membrane Proteins/chemistry , Membrane Proteins/genetics , Molecular Sequence Data , Mucins/metabolism , Protozoan Proteins/chemistry , Protozoan Proteins/genetics , Sequence Analysis, DNA , Trypanosoma cruzi/pathogenicity , Vero Cells
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