Luis Fernando Viana Furtado

Parasitismo por Ascaris lumbricoides e seus aspectos epidemiológicos em crianças do Estado do Maranhão

INTRODUCTION Intestinal parasitoses are a serious public health problem of worldwide nature. These disorders are correlated with poor basic sanitation conditions coupled with lack of basic hygiene notions, and are observed especially among children. Among these parasites, the nematode Ascaris lumbricoides stands out, with high incidence in Brazil and worldwide. The prevalence and intensity of infection by this pathogen were analyzed by means of a cross-sectional study among children living in the municipality of Tutóia, State of Maranhão, between July and December 2008. METHODS The study population consisted of children between one and twelve years of age, totaling 220 individuals. Fecal samples were collected in households in vials containing MIF preserving solution and were processed using the spontaneous sedimentation technique. A standard questionnaire was applied to each parent or guardian, and the results were used for descriptive analysis on the study sample. RESULTS The prevalence of A. lumbricoides was 53.6%. Analysis on the questionnaires revealed alarming results regarding the degree of unhealthy condition to which the population is subjected, in addition to its poor hygiene habits. CONCLUSIONS The prevalence rate found in this study is a clear reflection of the lack of basic sanitation in the region studied. Thus, a public policy for raising awareness and combating this disease is needed.

Benzimidazole resistance in helminths: From problem to diagnosis

Helminth parasites cause significant morbidity and mortality in endemic countries. Given the severity of symptoms that helminths may elicit in the host, intervention with prophylactic and therapeutic measures is imperative. Treatment with benzimidazoles is the most widely used means of combatting these parasites. However, widespread use of these drugs can select for drug-resistant parasite strains. In this review, we approach the problem of benzimidazole resistance in helminths in both humans and animals, focusing on the properties of the drug, the molecular mechanisms of drug resistance and how resistance is diagnosed.

Prevalência e aspectos epidemiológicos de enteroparasitoses na população geronte de Parnaíba, Estado do Piauí

INTRODUCTION: Since intestinal parasites pose public health problems, this work aimed to determine certain epidemiological aspects of these pathologies in elderly individuals from Parnaiba. METHODS: Using the method of Hoffman, 294 fecal samples from individuals 65 years-old or over were analyzed. Each participant responded to a questionnaire used for description analysis of the population. RESULTS: The prevalence of intestinal parasites was 40.5%. The questionnaires showed alarming results regarding the noxious conditions in which this population is inserted. CONCLUSIONS: High prevalence of intestinal parasites exists among the elderly population of Parnaiba, indicating an epidemiological state of some concern.

Standardization and application of the tetraprimer ARMS-PCR technique for screening of the E198A SNP in the β-tubulin gene of hookworm populations in Brazil

The tetraprimer ARMS-PCR technique is efficient for SNP detection and can be used to search for polymorphisms associated with drug resistance. However, the establishment of this methodology is not always straightforward because of the constraints on primer design due to the restrictions of the polymorphic regions. Here, we describe the standardization of the tetraprimer ARMS-PCR methodology for the detection of a SNP at codon 198 of the Ancylostoma caninum β-tubulin gene. This SNP is associated with resistance to albendazole in various nematodes. The methodology was used to screen 327 individuals from 6 different locations. No mutation was found in any of the samples. This methodology will be useful for screening for the E198A SNP in the β-tubulin gene of canine hookworms in a broader population to determine whether this SNP is associated with benzimidazole resistance in this species. The method could also be adapted for the analysis of other SNPs in other nematode species.

Molecular analysis of the F167Y SNP in the β-tubulin gene by screening genotypes of two Ancylostoma caninum populations.

Mutations in the β-tubulin isotype 1 gene at codons 167 (F167Y), 198 (E198A) and 200 (F200Y) have been associated with benzimidazole resistance in helminths. The F200Y polymorphism has previously been described for Ancylostom caninum; however, the F167Y polymorphism has not been investigated in members of the Ancylostomatidae family. The aim of this study was to screen for the F167Y polymorphism in A. caninum isolates recovered from naturally infected dogs in two Brazilian states. No mutation was observed at codon 167 in the 230 analyzed samples from the two populations; however, it is possible that this change may be present at a low frequency in other populations of the same species. These results highlight the importance of monitoring the genetic basis involved in the drug resistance process.

DNA barcoding of metacestodes found in the Guerlinguetus ingrami (Rodentia: Sciuridae) reveals the occurrence of Hydatigera taeniaeformis sensu stricto (Cyclophyllidea: Taeniidae) in the Americas

The existence of cryptic species in the genus Hydatigera, cyclophillid cestodes, mainly of felids, was recently described based on molecular studies of parasites from Asia, Europe and Africa. However, the occurrence of H. taeniaeformis sensu stricto (s.s.), the species more widely distributed and with a presumed specificity for murid rodents as intermediate hosts, has not been formally described in Americas. In the present study, during necropsy of an Ingrams squirrel specimen, Guerlinguetus ingrami, found dead in the municipality of Belo Horizonte, Minas Gerais state, Brazil, strobilocerci were found in the liver. The metacestodes were subjected to morphological and molecular studies. Sequences of the COI barcode region were obtained and used for phylogenetic analyses. The morphology and measures of the rostellar hooks were compatible with the ones described for H. taeniaeformis s.s. This identification was confirmed by a molecular phylogenetic approach (96.2-99.7% similarity with isolates of the parasite from Europe and Asia). This is the first molecular confirmation of the existence of H. taeniaeformis s.s. on the American continent. Moreover, the involvement of sciurid rodents in the transmission of H. taeniaeformis s.s. is discussed here as a probable case of parasite spillover.

Development of new microsatellites for the hookworm Ancylostoma caninum and analysis of genetic diversity in Brazilian populations

Considering the great efforts towards formulating a vaccine against hookworms, and the concerns about the spread of drug resistance through hookworm populations, it is justified to study the molecular diversity and population genetic structure of these nematodes. This work had the aim to develop microsatellite markers to investigate the genetic structure and the molecular diversity of Brazilian populations of Ancylostoma caninum. Seven microsatellites markers were successfully used to characterize five Brazilian populations. These findings may contribute to a better comprehension of the ecology, patterns of transmission, drug resistances and development of immunotherapeutic strategies in hookworms.

Standardization and application of a modified RFLP-PCR methodology for analysis of polymorphisms linked to treatment resistance in Ancylostoma braziliense

BackgroundSingle nucleotide polymorphisms (SNPs) in codons 167, 198 and 200 of the beta-tubulin isotype 1 gene are associated with benzimidazoles resistance in many helminths. Codon 167 mutation has never been described in hookworms; however, polymorphisms in codons 198 and 200 have been described for Ancylostoma caninum and Necator americanus. These mutations have never been investigated in Ancylostoma braziliense; therefore, it is not known if they are present in this species and whether they are correlated with treatment resistance. The RFLP-PCR technique has been used to analyze these polymorphisms in some nematodes, but depending on the species, these alterations do not create or eliminate any restriction enzyme cleavage site, making it impossible to use this technique. Here, we describe the standardization and application of a modified RFLP-PCR technique for detecting polymorphisms in individual A. braziliense worms recovered from naturally infected dogs in two Brazilian states.ResultsThe molecular techniques used were sensitive, specific, and easy to apply. To our knowledge, we report for the first time the presence of a polymorphism at codon 198 of the beta-tubulin gene of A. braziliense (1/81; 95% CI: 0–3.69%).ConclusionsIt is not known whether the presence of the mutation in codon 198 of the beta-tubulin gene of A. braziliense has importance for this parasite. However, based on studies of other helminths, it is possible that this polymorphism is directly related to the resistance to benzimidazoles. This may be a major concern, since this nematode has considerable relevance as a parasite of canids and felids and as one of the agents of cutaneous larva migrans in humans. Standardized methodologies will be useful for screening for polymorphisms in the beta-tubulin gene of canine hookworms in a broader population. The method could also be adapted for the analysis of other SNPs in other nematode species.

PCR-RFLP screening of polymorphisms associated with benzimidazole resistance in Necator americanus and Ascaris lumbricoides from different geographical regions in Brazil

Ascaris lumbricoides and Necator americanus are soil-transmitted parasites with global geographic distribution, and they represent some of the most common and neglected infections in the world. Periodic treatment with mass drug administration (MDA) in endemic areas is the recommended action put forth by the World Health Organization. However, MDA can cause the selection of subpopulations that possess the genetic ability to overcome the mechanism of drug action. In fact, beta-tubulin gene mutations (codons 167, 198 and 200) are correlated with benzimidazole resistance in nematodes of veterinary importance. It is possible that these SNPs also have strong correlation with treatment resistance in the human geohelminths A. lumbricoides, Trichuris trichiura and hookworms. Here, we aimed to investigate the presence of some of these canonical molecular markers associated with parasite resistance to benzimidazole in N. americanus and A. lumbricoides collected from six Brazilian states. Nested-PCR and PCR-RFLP were used to detect mutations at codons 167 and 198 in 601 individual eggs of A. lumbricoides collected from 62 human stool samples; however, no mutations were found. Codons 198 and 200 were tested in 552 N. americanus eggs collected from 48 patients using the same methodology, which presented a relative frequency of 1.4% and 1.1%, respectively. The presence of these SNPs in N. americanus eggs is an important finding, indicating that with high benzimidazole drug pressure there is potential for benzimidazole resistance to be selected in this hookworm. However, at these low frequencies it does not indicate that there is at present any benzimidazole resistance problem. This is the first systematic study performed in South America, and the study yielded a landscape of the genetic variants in the beta-tubulin gene and anthelmintic resistance to soil-transmitted parasites detected by a simple, rapid and affordable genotyping assay of individual eggs.

Fishborne Zoonotic Trematodes Transmitted by Melanoides tuberculata Snails, Peru

We investigated the transmission of the fishborne trematodes Centrocestus formosanus and Haplorchis pumilio by Melanoides tuberculata snails in Peru. We report on results of experimental, morphological, and molecular approaches and discuss the potential risk for future human cases, given the existence of food habits in the country involving the ingestion of raw fish.