Zoonotic Giardia duodenalis assemblage A in northern sloth from Brazilian Amazon

BACKGROUND The parasite Giardia duodenalis infects a wide range of vertebrate hosts, including domestic and wild animals as well as humans. Giardia is genotyped into eight assemblages (A-H). Zoonotic assemblages A and B have already been identified in humans and wild and domestic animals (non-human primates and cats) from Brazilian Amazon and in the world. Due to its zoonotic/zooanthroponotic nature, surveillance initiatives and the definition of Giardia assemblages are important in order to characterise the epidemiological scenario and to implement further control measures. OBJECTIVES Determine assemblages of G. duodenalis in sloths from the Brazilian Amazon Region. METHODS Faecal parasitological examination of sloths from Amazonas State. Polymerase chain reaction (PCR) targeting the beta giardin (BG), and genes from multilocus sequence typing (MLST) scheme, amplicon sequencing and phylogenetic analysis. FINDINGS Here, we identified, by microscopy, Giardia in two northern sloths (Bradypus tridactylus). These two samples were submitted to molecular assays and it was revealed that both were infected by G. duodenalis assemblage A. Phylogenetic analysis showed that they belong to assemblage A within sequences from humans and wild and domestic animals. CONCLUSION Therefore, besides showing, by the first time, the current presence of this parasite in sloths, our findings reveals that this wild animal species would be part of the zoonotic/zooanthroponotic scenario of this parasite in the Brazilian Amazon.

Occurrence and molecular characterization of Giardia duodenalis from naturally infected dogs in the municipality of Santa Maria, Rio Grande do Sul, Brazil

ABSTRACT: Giardiasis is an important and prevalent zoonosis in dogs and humans caused by Giardia spp. The close relationship between pets and humans has physical, emotional and social benefits. The dogs have an important role in Giardia duodenalis cycle and transmission. This study aimed to verify the occurrence of the parasite in dogs from Central Region, in Santa Maria, Rio Grande do Sul State, Brazil, from April to October 2018. Dog feces (230) were submitted to Faust coproparasitological and molecular analyses. The positive samples in the nested-PCR (-giardin gene) were sent for DNA sequencing and phylogenetic analyses (Neighbor-Joining). The occurrence of G. duodenalis, was 5.6% (13/230) and 4.3% (10/230) detected by coproparasitological technique and nested-PCR, respectively. There was no difference in the sensitivity of the tests used. From the faecal samples analyzed, there were no differences among the variables: diagnostic techniques, local, sex, and age of the animals (p>0.05). Only in the stool examination methodology a difference was observed between the ages (p 0.05). G. duodenalis assemblages were C and D, frequently reported in dogs. The close relationship between dogs and people may allow co-infections of circulating parasites in the population, including Giardia spp. and increasing the risk of transmission of zoonotic agents.

RESUMO: A giardíase é uma zoonose importante e prevalente em cães e humanos, sendo causada por Giardia spp. A estreita relação entre animais de estimação e seres humanos traz benefícios físicos, emocionais e sociais. Os cães têm um papel importante no ciclo e transmissão de Giardia duodenalis. Este estudo teve como objetivo verificar a ocorrência do parasita em cães da Região Central, em Santa Maria, RS, Brasil, de abril a outubro de 2018. As fezes de cães (230) foram submetidas a técnica coproparasitológica de Faust e análises moleculares. As amostras positivas no nested-PCR (gene -giardin) foram enviadas para sequenciamento de DNA e posterior análise filogenética (Neighbor-Joining). A ocorrência de G. duodenalis foi de 5,6% (13/230) e 4,3% (10/230) detectados pela técnica coproparasitológica e nested-PCR, respectivamente. Não houve diferença na sensibilidade dos testes utilizados. Das amostras fecais analisadas, não houve diferenças entre as variáveis: técnicas de diagnóstico, local, sexo e idade dos animais (p>0,05). Somente na metodologia de exame de fezes observou-se diferença entre as idades (p 0,05). As assemblages de G. duodenalis encontradas foram C e D, frequentemente relatadas em cães. A estreita relação entre cães e pessoas pode permitir co-infecções de parasitas circulantes na população, incluindo Giardia spp. e aumentando o risco de transmissão de agentes zoonóticos.

Giardiasis in urban and rural Amazonas, Brazil is driven by zoonotic and cosmopolitan A and B assemblages

BACKGROUND Giardia duodenalis is a protozoan parasite that infects humans and other mammals and causes giardiasis worldwide. Giardia is genotyped into eight assemblages (A-H), with assemblages A and B considered zoonotic. OBJECTIVES The aim of this study was to determine the assemblages of G. duodenalis from individuals living in rural and urban areas of the Amazonas State. METHODS 103 human faecal specimens microscopically positive for the presence of Giardia obtained from four municipalities in Amazonas and four animal faecal specimens were genotyped based on the sequences of two genes, triosephosphate isomerase (TPI) and β-giardin (BG). FINDINGS In humans, assemblage A was the most represented with the identification of sub-assemblages AI, AII and AIII based on BG and sub-assemblages AI and AII based on TPI. Similarly, there is a diversity of sub-assemblage B considering BG (B and BIII) and TPI (B, BIII and BIV). In addition, we characterised homogeneous and heterogeneous genotypes comprising assemblages/sub-assemblages A and B in individuals from urban and rural areas. Here, for the first time, it was genotyped Giardia that infects animals from the Brazilian Amazon region. We identified sub-assemblage AI in one Ateles paniscus and two Felis catus and sub-assemblage BIV in one Lagothrix cana. MAIN CONCLUSIONS Therefore, humans and animals from the urban and rural Amazon share Giardia genotypes belonging to assemblages A and B, which are found in cosmopolitan regions around the world.

Occurrence and molecular characterization of Giardia duodenalis from naturally infected dogs in the municipality of Santa Maria, Rio Grande do Sul, Brazil / Ocorrência e caracterização molecular de Giardia duodenalis em cães naturalmente infectados no município de Santa Maria, Rio Grande do Sul, Brasil

Giardiasis is an important and prevalent zoonosis in dogs and humans caused by Giardia spp. The close relationship between pets and humans has physical, emotional and social benefits. The dogs have an important role in Giardia duodenalis cycle and transmission. This study aimed to verify the occurrence of the parasite in dogs from Central Region, in Santa Maria, Rio Grande do Sul State, Brazil, from April to October 2018. Dog feces (230) were submitted to Faust coproparasitological and molecular analyses. The positive samples in the nested-PCR (β-giardin gene) were sent for DNA sequencing and phylogenetic analyses (Neighbor-Joining). The occurrence of G. duodenalis, was 5.6% (13/230) and 4.3% (10/230) detected by coproparasitological technique and nested-PCR, respectively. There was no difference in the sensitivity of the tests used. From the faecal samples analyzed, there were no differences among the variables: diagnostic techniques, local, sex, and age of the animals (p>0.05). Only in the stool examination methodology a difference was observed between the ages (p<0.05). G. duodenalis assemblages were C and D, frequently reported in dogs. The close relationship between dogs and people may allow co-infections of circulating parasites in the population, including Giardia spp. and increasing the risk of transmission of zoonotic agents.(AU)

A giardíase é uma zoonose importante e prevalente em cães e humanos, sendo causada por Giardia spp. A estreita relação entre animais de estimação e seres humanos traz benefícios físicos, emocionais e sociais. Os cães têm um papel importante no ciclo e transmissão de Giardia duodenalis. Este estudo teve como objetivo verificar a ocorrência do parasita em cães da Região Central, em Santa Maria, RS, Brasil, de abril a outubro de 2018. As fezes de cães (230) foram submetidas a técnica coproparasitológica de Faust e análises moleculares. As amostras positivas no nested-PCR (gene β-giardin) foram enviadas para sequenciamento de DNA e posterior análise filogenética (Neighbor-Joining). A ocorrência de G. duodenalis foi de 5,6% (13/230) e 4,3% (10/230) detectados pela técnica coproparasitológica e nested-PCR, respectivamente. Não houve diferença na sensibilidade dos testes utilizados. Das amostras fecais analisadas, não houve diferenças entre as variáveis: técnicas de diagnóstico, local, sexo e idade dos animais (p>0,05). Somente na metodologia de exame de fezes observou-se diferença entre as idades (p<0,05). As assemblages de G. duodenalis encontradas foram C e D, frequentemente relatadas em cães. A estreita relação entre cães e pessoas pode permitir co-infecções de parasitas circulantes na população, incluindo Giardia spp. e aumentando o risco de transmissão de agentes zoonóticos.(AU)

Ultrastructural and proapoptotic-like effects of kaempferol in Giardia duodenalis trophozoites and bioinformatics prediction of its potential protein target

BACKGROUND Kaempferol (KPF) is a flavonoid with antiparasitic activity including experimental giardiasis which mechanism of action is unknown. OBJECTIVE To analyse the cytotoxic effects of KPF on Giardia duodenalis trophozoites and to identify a likely parasite target of this compound. METHODS We used inhibitory concentrations of KPF (IC25, IC50 and IC100) and albendazole (ABZ) as reference drug. The ultrastructure of the trophozoites was analysed by transmission electron microscopy (TEM) whilst apoptosis/necrosis, production of reactive oxygen species (ROS) and cell cycle progression were assessed by flow cytometry (FCM) and confocal laser microscopy (CLM). Ligand-protein docking analyses were carried out using KPF structure from a drug library and crystal structure of a G. duodenalis aldose reductase (GdAldRed) homolog. RESULTS KPF provoked appearance of perinuclear and periplasmic spaces devoid of cytosolic content and multilamellar structures. KPF induced proapoptotic death associated with partial arrest in the S phase without ROS production. Bioinformatics approaches predicted that GdAldRed is a viable KPF target (ΔG = -7.09 kCal/mol), exhibiting 92% structural identity and a similar coupling pattern as its human homolog. CONCLUSIONS KPF exerted a proapoptotic effect on G. duodenalis trophozoites involving partial interruption of DNA synthesis without oxidative stress or structure damage to chromatin and cytoskeletal structures. GdAldRed is a likely target underlying its antigiardial activity.

In vitro-induction of metronidazole-resistant Giardia duodenalis is not associated with nucleotide alterations in the genes involved in pro-drug activation

Giardiasis is an infectious disease caused by Giardia duodenalis. The pro-drug metronidazole (MTZ) is the first-line treatment for giardiasis. Parasite's proteins as pyruvate:ferredoxin oxidoreductase (PFOR), ferredoxin (Fd), nitroreductase-1 (NR-1) and thioredoxin reductase (TrxR) participate in MTZ activation. Here, we showed Giardia trophozoites long-term exposed to MTZ presented higher IC50 than controls, showing the drug influenced the parasite survival. That reduction in MTZ's susceptibility does not seem to be related to mutations in the genes pfor, fd, nr-1 or trxr. It points that different mechanism as alterations in other metabolic pathways can account for Giardia resistance to MTZ therapy.

Epidemiology of Giardia duodenalis assemblages in Brazil: there is still a long way to go

Giardia duodenalis infection is distributed worldwide and can achieve prevalence around 60%, especially in developing countries. This protozoan is divided into eight assemblages, in which A and B have high zoonotic potential, whereas C to H are host-specific. This scenario is changing as molecular studies progress, highlighting that knowledge on host-specificity still has a long way to go. Understanding the players involved in transmission routes enables rational designs of control strategies. Considering the high prevalence of giardiasis, this review aims to gather together the data on available studies on the distribution of G. duodenalis assemblages in Brazil until September 2020.

Molecular Prevalence and Genotypes of Cryptosporidium parvum and Giardia duodenalis in Patients with Acute Diarrhea in Korea, 2013–2016

Cryptosporidium parvum and Giardia duodenalis are the main diarrhea-causing parasitic pathogens; however, their prevalence in Korea is unknown. Here, we conducted a survey to determine the prevalence and genotype distribution of these 2 pathogens causing acute diarrhea in 8,571 patients hospitalized in 17 Regional Institute of Health Environment sites in Korea, during 2013–2016. C. parvum and G. duodenalis were detected and genotyped by nested PCR, and the isolate were molecularly characterized by sequencing the glycoprotein 60 (Gp60) and β-giardin genes, respectively. The overall prevalence of C. parvum and G. duodenalis was 0.37% (n=32) and 0.55% (n=47), respectively, and both pathogens were more prevalent in children under 9 years old. Molecular epidemiological analysis showed that the C. parvum isolates belonged to the IIa family and were subtyped as IIaA13G2R1, IIaA14G2R1, IIaA15G2R1, and IIaA18G3R1. Analysis of the β-giardin gene fragment from G. duodenalis showed that all positive strains belong to assemblage A. This is the first report on the molecular epidemiology and subtyping of C. parvum and G. duodenalis in such a large number of diarrheal patients in Korea. These results highlight the need for continuous monitoring of these zoonotic pathogens and provide a basis for implementing control and prevention strategies. Further, the results might be useful for epidemiological investigation of the source of outbreak.

Single dose secnidazole treatment efficiency against naturally occuring Giardia duodenalis infection in calves / Eficacia del tratamiento con secnidazol en dosis única contra la infección natural de Giardia duodenalis en terneros

Abstract Objective. In the present study the aim was to establish the efficacy of 30 mg/kg single dose secnidazol in calves naturally infected with Giardia duodenalis. Materials and methods. In an attempt to perform original study a total of 18 calves, from various breed, age and of both sexes were enrolled. Diagnosis was based on detection of trophozoit and/or cysts on fecal flotation among calves naturally infected with G. duodenalis, besides by use of rapid diagnostic test kits working with solid phase immunochromatographic principles and β-giardin nested- Polymerase Chain Reaction aplication. Cyst count per gram of feces were performed among days 0, 3, 7 and 10 in all cases. On days 0 and 10 hematological (WBC, RBC, HCT, MCHC, PLT) and serum biochemical (ALT, AST, creatinine, urea) values were determined. Results. Two different groups of calves composed of secnidazole group (n:9) and control group (n:9) were enrolled. Among calves enrolled in treatment group secnidazole was administered at a single dosage of 30 mg/kg perorally on day 0, whereas control group were left without any active ingredient. Cyst count per gram feces, hematological and serum biochemical values were analyzed among groups and intragroup comparisons. Giardia duodenalis assemblage A3 was detected in all 18 calves. On days 3, 7, and 10 there was significant (p˂0.001) reduction in cyst excretion; whereas evaluation of mean geometric cyst excretion revealed 100% reduction on days 7 and 10. Among two group statistical analysis of hematological and serumbiochemical variables revealed no statistical significance on days 0 and 10. Conclusions. In conclusion secnidazole at a single dose of 30 mg/kg might be practically appliable, reasonably priced, safety, completely effective and causing rapid recovery treatment protocole for therapy of calves with Giardiasis.

Resumen Objetivo. En el presente estudio se pretendía establecer la eficacia de 30 mg / kg de dosis única de secnidazol en terneros naturalmente infectados con Giardia duodenalis. Materiales y métodos. En un intento de realizar un estudio original, se matricularon 18 terneros, de distintas raza, edad y de ambos sexos. El diagnóstico se basó en la identificación de trofozoítos y /o quistes en la flotación fecal entre terneros naturalmente infectados con G. duodenalis, además de utilizar kits de diagnóstico rápido que funcionan con principios inmunocromatográficos en fase sólida y aplicación de Reacción en Cadena de Polimerasa anidada con β-giardina. El conteo de quistes por gramo de heces se realizó entre los días 0, 3, 7 y 10 en todos los casos. En los días 0 y 10 se determinaron los valores hematológicos (WBC, RBC, HCT, MCHC, PLT) y suero bioquímico (ALT, AST, creatinina, urea). Resultados. Se inscribieron dos grupos diferentes de terneros tratados con secnidazol (n: 9) y el grupo control (n: 9). Entre los becerros incluidos en el grupo de tratamiento se administró secnidazol en una dosis única de 30 mg/kg oralmente al día 0, mientras que el grupo de control se dejó sin ningún ingrediente activo. Se analizó el conteo de quistes por gramo de heces, valores hematológicos y bioquímicos de suero entre grupos y comparaciones intragrupo. Giardia duodenalis ensamblaje A3 se identificó en los 18 terneros. En los días 3, 7 y 10 hubo una reducción significativa (p˂0.001) en la excreción de quistes; Mientras que la evaluación de la excreción geométrica promedio de quistes reveló una reducción del 100% en los días 7 y 10. Entre los dos grupos de análisis estadístico de las variables hematológicas y sero-bioquímicas no reveló significación estadística en los días 0 y 10. Conclusión. En conclusión secnidazol en una dosis única de 30 mg/kg podría ser prácticamente aplicable, a un precio razonable, la seguridad, completamente eficaz y causando un tratamiento de recuperación rápida protocolo para la terapia de terneros con Giardiasis.

Modification of the Alere GIARDIA Ag TEST immunochromatography KIT methodology for its use in frozen fecal sediment of dogs and cats

ABSTRACT Giardia duodenalis is a worldwide intestinal parasite and is one of the most frequent protozoa species infecting dogs and cats. This study aimed to modify the methodology of Alere GIARDIA Ag TEST KIT for its use in frozen fecal sediments with different storage times in a freezer (-20°C), thus expanding the range of use of this methodology. One hundred fecal sediments from dogs (n=50) and cats (n=50) previously examined by optical microscopy for Giardia cysts were selected for this study. The agreement between the modified immunochromatography and microscopy results was calculated by Kappa coefficient. To evaluate the performance of the modified immunochromatography assay on samples with different storage time, the fecal sediments were divided into three groups according to the time of storage in a freezer: (a) ≤ 1 year (n=37); (b) > 1 year and ≤ 3 years (n=39); (c) > 10 years (max. 13 years) (n=24). The results obtained by the modified immunochromatography assay demonstrates a higher sensitivity of this technique when compared with microscopy, regardless of the frozen storage time. These results allow for the use of this methodology in a greater scope of analysis, especially in frozen fecal sediment triage in sample collections, enabling epidemiological and comparative analysis along different decades.

Intestinal parasite prevalence in schoolchildren from northwestern Rio Grande do Sul state, Brazil

Parasites are often a public health hazard, especially among schoolchildren. The transmission of these parasites is associated with poor hygiene and sanitation, as well as close contact between people. This study aimed to evaluate the occurrence of intestinal parasitic infections in schoolchildren from the rural and urban areas of the Palmeira das Missões county, a major town in northwestern Rio Grande do Sul State. Fecal samples from 209 schoolchildren aged three to fourteen were analyzed. The Hoffman, Pons and Janer, Faust, Ritchie, and Kinyoun's stain for coccidea parasite detection techniques were applied. 59.3% of the patients were positive for pathogenic or commensal parasites. The most common parasites were Ascaris lumbricoides, Entamoeba coli, Giardia duodenalis and Cryptosporidium spp., respectively. There were no significant differences in positivity among schoolchildren from rural and urban areas. Low family income and the mothers' schooling (up to 8 years of education) were statistically significant for the presence of parasites. This is the first parasite study carried out in the Palmeira das Missões county

Detection of zoonotic and livestock-specific assemblages of Giardia duodenalis in free-living wild lizards / Detecção de genótipos de Giardia duodenalis zoonóticos e específicos de ruminantes domésticos em lagartos selvagens

Abstract Giardia duodenalis is a zoonotic parasite that infects the gut of a wide range of vertebrates, including numerous wildlife species. However, little is known about this protozoan parasite in reptiles. Fecal samples from 31 wild lizards were collected in Galicia (northwest Spain) and screened for the presence of Giardia by PCR amplification and sequencing of the ITS1-5.8S-ITS2 region in the ribosomal unit. This allowed detection of the parasite in 5 samples (16.1%), and enabled identification of G. duodenalis assemblage A2 in two samples of Iberian rock lizard (Iberolacerta monticola), G. duodenalis assemblage B in other two samples of I. monticola, and G. duodenalis assemblage E in one sample of Bocage's wall lizard (Podarcis bocagei). The results obtained after PCR amplification and sequencing of the SSU-rDNA gene confirmed the presence of G. duodenalis assemblage A in two samples of I. monticola. This is the first report of G. duodenalis in free-living lizards, although further studies are needed to distinguish between actual infection and mechanical dissemination of cysts. The detection of zoonotic and livestock-specific assemblages of G. duodenalis demonstrates the wide environmental contamination by this parasite, possibly due to human activities.

Resumo Giardia duodenalis é um parasito zoonótico que infecta o intestino delgado de uma ampla gama de vertebrados, sendo detectado em numerosas espécies selvagens. No entanto, pouco se conhece sobre a presença deste parasito protozoário em répteis. Para estudar a presença de Giardia, foram obtidas amostras fecais provenientes de 31 lagartos e coletadas em diferentes localizações de Galicia (Noroeste da Espanha). Mediante a aplicação da técnica de PCR e posterior sequenciamento da região ITS1-5.8S-ITS2 da unidade ribossômica, detectou-se Giardia em 5 amostras (16,1%), identificando-se o genótipo A2 de G. duodenalis em 2 amostras de lagartos da montanha (Iberolacerta monticola), G. duodenalis genótipo B em outras 2 amostras de I. monticola e G. duodenalis genótipo E em outra amostra de lagarto de Bocage (Podarcis bocagei). Os resultados obtidos, após amplificação e sequenciamento de um fragmento do gene SSU-rDNA, confirmam a presença de G. duodenalis genótipo A em 2 amostras de I. monticola. Esta é a primeira vez que se descreve G. duodenalis em lagartos selvagens, embora sejam necessários outros estudos complementares para confirmar se estes animais sofrem uma infecção real ou se apenas atuam como disseminadores mecânicos da contaminação ambiental. Além disso, a detecção de genótipos zoonóticos e específicos de ruminantes domésticos demonstra a contaminação do ambiente selvagem por G. duodenalis, possivelmente devido à atividade humana.

Giardia duodenalis infection among rural communities in Yemen: A community-based assessment of the prevalence and associated risk factors

Objective To investigate the prevalence and risk factors of Giardia infection among rural communities among rural communities in Yemen. Methods In this cross-sectional study, a total of 605 stool samples were collected and screened for the presence of Giardia duodenalis (G. duodenalis) cysts and/or trophozoites by using three different diagnostic methods: direct smear, formalin-ether sedimentation, and trichrome staining. A pre-tested questionnaire was used to collect information on the demographic, socioeconomic, behavioural and environmental characteristics of the participants. Results Overall, 28.1% (170/605) of the participants were infected by G. duodenalis. The prevalence was significantly higher among male participants compared to female (P = 0.034); however, it was not significant among different age groups (P > 0.05). Univariate and multivariate analyses identified four variables as the significant key risk factors of Giardia infection among the sampled communities. These are, in addition to being of the male gender, using unsafe water sources for drinking water, not washing hands after defecation, presence of other family members infected with Giardia, and close contact with domestic animals. Conclusions The study reveals that Giardia infection is still prevalent among rural communities in Yemen. The provision of clean and safe drinking water, proper sanitation, and health education regarding personal hygiene practices, particularly handwashing, as well as identifying and treating infected family members is imperative and these interventions should be considered in a strategy to control intestinal parasites among these communities in order to curtail the transmission and morbidity caused by G. duodenalis.

Giardia duodenalis infection among rural communities in Yemen: A community-based assessment of the prevalence and associated risk factors

OBJECTIVE@#To investigate the prevalence and risk factors of Giardia infection among rural communities among rural communities in Yemen.@*METHODS@#In this cross-sectional study, a total of 605 stool samples were collected and screened for the presence of Giardia duodenalis (G. duodenalis) cysts and/or trophozoites by using three different diagnostic methods: direct smear, formalin-ether sedimentation, and trichrome staining. A pre-tested questionnaire was used to collect information on the demographic, socioeconomic, behavioural and environmental characteristics of the participants.@*RESULTS@#Overall, 28.1% (170/605) of the participants were infected by G. duodenalis. The prevalence was significantly higher among male participants compared to female (P = 0.034); however, it was not significant among different age groups (P > 0.05). Univariate and multivariate analyses identified four variables as the significant key risk factors of Giardia infection among the sampled communities. These are, in addition to being of the male gender, using unsafe water sources for drinking water, not washing hands after defecation, presence of other family members infected with Giardia, and close contact with domestic animals.@*CONCLUSIONS@#The study reveals that Giardia infection is still prevalent among rural communities in Yemen. The provision of clean and safe drinking water, proper sanitation, and health education regarding personal hygiene practices, particularly handwashing, as well as identifying and treating infected family members is imperative and these interventions should be considered in a strategy to control intestinal parasites among these communities in order to curtail the transmission and morbidity caused by G. duodenalis.

High Frequency of Enteric Protozoan, Viral, and Bacterial Potential Pathogens in Community-Acquired Acute Diarrheal Episodes: Evidence Based on Results of Luminex Gastrointestinal Pathogen Panel Assay

Infectious diarrhea is endemic in most developing countries. We aimed to investigate the protozoan, viral, and bacterial causes of acute diarrhea in Taif, Saudi Arabia. A cross-sectional prospective 1-year study was conducted on 163 diarrheal patients of various ages. Stool samples were collected, 1 per patient, and tested for 3 protozoa, 3 viruses, and 9 bacteria with the Luminex Gastrointestinal Pathogen Panel. Overall, 53.4% (87/163) of samples were positives (20.8% protozoa, 19.6% viruses, 2.8% bacteria, and 9.8% mixed). Rotavirus (19.6%), Giardia duodenalis (16.5%), and Cryptosporidium spp. (8.5%) were the mostly detected pathogens. Adenovirus 40/41 (4.2%), Salmonella (3%), Shiga toxin-producing Escherichia coli (3%), and Entamoeba histolytica (2.4%) were also detected. Norovirus GI/II, Vibrio cholerae, Yersinia enterocolitica, and Clostridium difficile toxin A/B were not detected in any patients. All pathogens were involved in coinfections except E. histolytica. Giardia (5.5%) and rotavirus (3%) were the most commonly detected in co-infections. Enterotoxigenic E. coli (2.4%), Campylobacter spp. (2.4%), E. coli 0157 (1.8%), and Shigella spp. (1.2%) were detected in patients only as co-infections. Infections were more in children 0–4 years, less in adults 40 years, with statistically significant differences in risk across age groups observed with rotavirus (P < 0.001), Giardia (P=0.006), and Cryptosporidium (P=0.036) infections. Lastly, infections were not significantly more in the spring. This report demonstrates the high burden of various enteropathogens in the setting. Further studies are needed to define the impact of these findings on the clinical course of the disease.

The efficacy of chloroquine treatment against naturally occuring Giardia duodenalis infection in lambs / La eficacia del tratamiento de cloroquina contra la infección natural por Giardia duodenalis en corderos

ABSTRACT Objective. The aim of the present study was to determine the efficacy of Chloroquine (Cq), an antimalarial medicine, administered at a dose of 2.5 mg/kg, orally, during 5 consecutive days, in Sakiz and Merino lambs naturally infected with Giardia duodenalis. Materials and methods. To this extent weaned 10 weeks of aged lambs were enrolled and randomly assigned into two groups based on treatment (group C, n=18 lambs treated with Cq) and placebo (group P, n=8 untreated control animals). Diagnosis was based on detection of trophozoit and/or cysts on fecal flotation. Cyst count per gram feces (days 0, 3, 7 and 10) was analyzed among groups. Results. During the trial, regarding the efficacy of Cq on days 3., 7., and 10. There was significant (p<0.01) reduction in cyst excretion; whereas evaluation of mean geometric cyst excretion revealed 100% reduction. Conclusions. There was a very high (100%) reduction in cyst excretion in the Cq treatment group compared to the positive untreated control group on days 3, 7 and 10, resulting in a significant (p<0.001) reduction, making Cq, safety, and thus highly effective treatment option of lambs with naturally occuring giardiasis.

RESUMEN Objetivo . El objetivo del presente estudio fue determinar la eficacia de la cloroquina, un medicamento contra la malaria, administrado a una dosis de 2.5 mg/kg, por vía oral, durante 5 días consecutivos, en Corderos Sakiz y Merino infectados naturalmente con Giardia duodenalis. Materiales y metodos. En esta medida destetados 10 semanas de edad fueron incluidos y asignados al azar en dos grupos sobre la base del tratamiento (grupo C, n = 18 corderos tratados con cloroquina) y placebo (grupo P, n = 8 animales de control no tratados). El diagnóstico se basa en la detección de trophozoit y/o quistes en la flotación fecal. Quiste recuento por gramo de heces (día 0, 3, 7 y 10) se analizó entre los grupos. Resultados. Respecto a la eficacia de la cloroquina en los días 3, 7 y 10 existió una reducción significativa (p<0.01) en la excreción de quistes; mientras que la evaluación de la media de la excreción de quistes reveló una reducción del 100%. Conclusiones. Durante el estudio hubo una reducción del 100% en la excreción de quistes en el grupo de tratamiento con cloroquina en comparación con el grupo control no tratado positivo en los días 3, 7 y 10, lo que resulta en una disminución significativa (p<0.001) por lo que la cloroquina razonablemente es una opción de tratamiento por el costo, seguridad, y por lo tanto muy eficaz de corderos infectados naturalmente con giardiasis.

Avaliação do consumo de Lactobacillus casei shirota para o controle de giardíase em crianças / Evaluation of Lactobacillus casei shirota consumption for giardiasis control in children

A giardíase é uma importante doença parasitária de elevada ocorrência mundial e, apesar de comumente ser considerada de bom prognóstico, sabe-se que crianças e pacientes imunocomprometidos podem apresentar perda de peso e depauperamento físico e mental. Há um grande número de drogas com atividade giardicida, mas, no decorrer dos anos, alguns protozoários têm apresentado resistência à maioria dos tratamentos recomendados, o que exige o desenvolvimento de novas estratégias para o controle de Giardia duodenalis. No presente trabalho, objetivou-se avaliar a eficácia do uso de Lactobacillus casei Shirota para o controle da giardíase em crianças naturalmente infectadas. Para tanto, foram selecionadas 12 crianças com giardíase clínica e laboratorialmente diagnosticada, das quais 6 compuseram o grupo controle, sem tratamento, e as demais consumiram 1,6x1010 lactobacilos vivos, diariamente, da espécie Lactobacillus casei Shirota, mediante ingestão de uma bebida láctea comercialmente disponível. Depois do início do tratamento, foi determinado o número de cistos eliminados por meio de exame coproparasitológico, segundo o método de Faust com modificações, e observação da consistência das amostras fecais coletadas. Transcorridos 21 dias de tratamento, não houve detecção de formas evolutivas de Giardia duodenalis nas fezes dos indivíduos que consumiram a bebida láctea e verificou-se melhora na consistência fecal, o que não ocorreu com o grupo controle, demonstrando-se, assim, o potencial de L. casei Shirota para o controle de giardíase em crianças.

Evidence of heterozygosity and recombinant alleles in single cysts of Giardia duodenalis / Evidência de heterozigose e alelos recombinantes em cistos individualizados de Giardia duodenalis

Abstract Giardia duodenalis is divided into eight assemblages (named A to H). Isolates of assemblage A are divided into four sub-assemblages (AI, AII, AIII and AIV). While isolates of sub-assemblage AII are almost exclusively detected in human hosts, isolates of assemblage B are encountered in a multitude of animal hosts and humans. Here, we isolated single cysts of G. duodenalis from a human stool sample and found that one of them had overlaps of assemblage AII and B alleles and an unexpectedly high number of variants of the beta-giardin (Bg) and GLORF-C4 (OrfC4) alleles. In addition, one of the Bg alleles of that cyst had a fragment of sub-assemblage AII interspersed with fragments of assemblage B, thus indicating that this allele may be a recombinant between sequences A and B. Our results are unprecedented and put a check on the statement that different assemblages of G. duodenalis represent species with different host specificities.

Resumo A espécie Giardia duodenalis é dividida em oito grupos (nomeados de A a H). Isolados do grupo A são divididos em quatro subgrupos (AI, AII, AIII and AIV). Enquanto isolados do subgrupo AII são detectados quase exclusivamente em hospedeiros humanos, isolados do subgrupo B são encontrados em uma grande variedade de hospedeiros entre animais e humanos. Neste trabalho, foi constatado que, dentre diversos cistos individualizados de G. duodenalis provenientes de fezes de origem humana, um cisto continha os alelos AII e B e um número inesperado de variantes de alelos codificadores de beta giardina e GLORF-C4. Ainda, um dos alelos beta giardina desse cisto possuía fragmentos AII intercalando um fragmento B, indicando que esse alelo pode ser um recombinante entre alelos AII e B. Os resultados aqui apresentados são inéditos e colocam em dúvida o conceito atual de que os diferentes grupos de G. duodenalis representam espécies distintas com diferentes graus de especificidade por hospedeiros.

Comparação de seis métodos de extração de DNA genômico de Giardia duodenalis / Comparison of six methods for genomic DNA extraction from Giardia duodenalis CYSTS

Introdução: Giardia duodenalis é um parasito gastrintestinal que infecta o homem e uma variedade de animais domésticos e silvestres. A G. duodenalis em amostras de origem fecal humana pertence a dois principais grupos genéticos, as assemblages A e B, também encontradas em animais. Amostras fecais foram analisadas e a extração e purificação do DNA foram realizadas para avaliar a eficácia da reação em cadeia da polimerase (PCR) na detecção do gene gdh de cistos de G. duodenalis. Métodos: A obtenção de amostras de DNA adequadas para protocolos de amplificação genotípicas foi feita por meio da PCR, comparando seis métodos de extração de DNA de cistos de G. duodenalis em amostras purificadas e não purificadas. Foram realizados os métodos QIAmp DNA Stool Mini Kit, processo de congelamento e descongelamento, ultrassom, uso de glass beads, desnaturação com formamida e o método convencional (fenol/clorofórmio). Resultados: Os métodos com ultrassom e uso de glass beads foram mais efetivos na extração do DNA. Não houve diferença entre o uso de amostras purificadas e não purificados para extração de DNA do protozoário. Conclusão: Verificou-se que tanto amostras purificadas como não purificadas podem ser usadas para extração de DNA de G. Duodenalis. Embora vários métodos de extração de DNA sejam preconizados na literatura, no presente estudo o uso de ultrassom e glass beads foram mais eficazes.

Introduction: Giardia duodenalis is a gastrointestinal parasite that infects human beings and a wide range of domestic and wild animal species. In humans, G.duodenalis samples belong to two major genetic groups, assemblages A and B, also found in animals. Human fecal samples have been approved to analyze the effects of DNA purification and extraction to assess the effectiveness of Polymerase Chain Reaction (PCR) in detecting the gene gdh from G. duodenalis cysts. Methods: Suitable DNA samples for genotypic amplification protocols were obtained by means of PCR, comparing six methods for DNA extraction from G. duodenalis cysts in purified and unpurified samples: QIAmp DNA Stool Mini kit, freeze-thaw procedure, sonication glass bead disruption, formamide denaturation, and conventional method (phenol/chloroform). Results: The methods of sonication and the use of glass beads were more effective in extracting DNA. There was no difference between the use of purified and unpurified samples for protozoan DNA extraction. Conclusions: In the present study we verified that both purified and unpurified samples can be used to G. duodenalis DNA extraction.Though various DNA extraction methods are recommended in the literature, the use of sonicator and glass beads were more effective in this study.

Occurrence and Molecular Identification of Giardia duodenalis from Stray Cats in Guangzhou, Southern China

The objective of this study was to genetically characterize isolates of Giardia duodenalis and to determine if zoonotic potential of G. duodenalis could be found in stray cats from urban and suburban environments in Guangzhou, China. Among 102 fresh fecal samples of stray cats, 30 samples were collected in Baiyun district (urban) and 72 in Conghua district (suburban). G. duodenalis specimens were examined using light microscopy, then the positive specimens were subjected to PCR amplification and subsequent sequencing at 4 loci such as glutamate dehydrogenase (gdh), triose phosphate isomerase (tpi), beta-giardin (bg), and small subunit ribosomal RNA (18S rRNA) genes. The phylogenetic trees were constructed using obtained sequences by MEGA5.2 software. Results show that 9.8% (10/102) feline fecal samples were found to be positive by microscopy, 10% (3/30) in Baiyun district and 9.7% (7/72) in Conghua district. Among the 10 positive samples, 9 were single infection (8 isolates, assemblage A; 1 isolate, assemblage F) and 1 sample was mixed infection with assemblages A and C. Based on tpi, gdh, and bg genes, all sequences of assemblage A showed complete homology with AI except for 1 isolate (CHC83). These findings not only confirmed the occurrence of G. duodenalis in stray cats, but also showed that zoonotic assemblage A was found for the first time in stray cats living in urban and suburban environments in China.