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Giardiasis is an intestinal infection that affects more than two hundred million people annually worldwide; it is caused by the flagellated protozoan Giardia duodenalis. In tropical countries and in low or middle-income settings, like Brazil, its prevalence can be high. There is currently no systematic review on the presence of G. duodenalis in patients, animals or water sources in Brazil. This systematic review was performed according to recommendations established by Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA). As databases for our searches, we have used PubMed, Embase, Scopus and the Brazilian database SciELO using the keywords «Giardia*» and «Brazil». This systematic review identified research studies related to G. duodenalis in water, giardiasis in animals, prevalence of giardiasis across Brazilian regions, genotyping of strains isolated in humans, and giardiasis in indigenous populations. We also propose a network of G. duodenalis transmission in Brazil based on genotypes analyses. This is the first time within the last twenty years that a review is being published on the occurrence of G. duodenalis in Brazil, addressing relevant issues such as prevalence, molecular epidemiology and analytical methods for parasite detection.

Giardia duodenalis is a flagellate protozoan that parasitizes humans and several other mammals. Protozoan contamination has been regularly documented at important environmental sites, although most of these studies were performed at the species level. There is a lack of studies that correlate environmental contamination and clinical infections in the same region. The aim of this study is to evaluate the genetic diversity of a set of clinical and environmental samples and to use the obtained data to characterize the genetic profile of the distribution of G. duodenalis and the potential for zoonotic transmission in a metropolitan region of Brazil. The genetic assemblages and subtypes of G. duodenalis isolates obtained from hospitals, a veterinary clinic, a day-care center and important environmental sites were determined via multilocus sequence-based genotyping using three unlinked gene loci. Cysts of Giardia were detected at all of the environmental sites. Mixed assemblages were detected in 25% of the total samples, and an elevated number of haplotypes was identified. The main haplotypes were shared among the groups, and new subtypes were identified at all loci. Ten multilocus genotypes were identified: 7 for assemblage A and 3 for assemblage B. There is persistent G. duodenalis contamination at important environmental sites in the city. The identified mixed assemblages likely represent mixed infections, suggesting high endemicity of Giardia in these hosts. Most Giardia isolates obtained in this study displayed zoonotic potential. The high degree of genetic diversity in the isolates obtained from both clinical and environmental samples suggests that multiple sources of infection are likely responsible for the detected contamination events. The finding that many multilocus genotypes (MLGs) and haplotypes are shared by different groups suggests that these sources of infection may be related and indicates that there is a notable risk of human infection caused by Giardia in this region.

BACKGROUND: Several species of protozoa cause acute or chronic gastroenteritis in humans, worldwide. The burden of disease is particularly high among children living in developing areas of the world, where transmission is favored by lower hygienic standards and scarce availability of safe water. However, asymptomatic infection and polyparasitism are also commonly observed in poor settings. Here, we investigated the prevalence of intestinal protozoa in two small fishing villages, Porto Said (PS) and Santa Maria da Serra (SM), situated along the river Tietê in the State of São Paolo, Brazil. The villages lack basic public infrastructure and services, such as roads, public water supply, electricity and public health services. METHODS: Multiple fecal samples were collected from 88 individuals in PS and from 38 individuals in SM, who were asymptomatic at the time of sampling and had no recent history of diarrheal disease. To gain insights into potential transmission routes, 49 dog fecal samples (38 from PS and 11 from SM) and 28 river water samples were also collected. All samples were tested by microscopy and PCR was used to genotype Giardia duodenalis, Blastocystis sp., Dientamoeba fragilis and Cryptosporidium spp. RESULTS: By molecular methods, the most common human parasite was Blastocystis sp. (prevalence, 45% in PS and 71% in SM), followed by D. fragilis (13.6% in PS, and 18.4% in SM) and G. duodenalis (18.2% in PS and 7.9% in SM); Cryptosporidium spp. were not detected. Sequence analysis revealed large genetic variation among Blastocystis samples, with subtypes (STs) 1 and 3 being predominant, and with the notable absence of ST4. Among G. duodenalis samples, assemblages A and B were detected in humans, whereas assemblages A, C and D were found in dogs. Finally, all D. fragilis samples from humans were genotype 1. A single dog was found infected with Cryptosporidium canis. River water samples were negative for the investigated parasites. CONCLUSIONS: This study showed a high carriage of intestinal parasites in asymptomatic individuals from two poor Brazilian villages, and highlighted a large genetic variability of Blastocystis spp. and G. duodenalis.

Giardia and Cryptosporidium have caused numerous outbreaks of diarrhea as a result of the ingestion of water contaminated with sewage. In Brazil, the efficiency of Giardia and Cryptosporidium removal by combined fixed-film systems has rarely been studied. The aims of the present study were therefore to verify the removal efficiency of Giardia and Cryptosporidium by a combined system (anaerobic/anoxic filter and aerated submerged biofilter) and to perform the genetic characterization of these parasites. The (oo)cysts were detected by centrifuge concentration and membrane filtration from raw sewage, effluents, adhered biomass, and sludge samples. Immunofluorescence assay and differential interference contrast microscopy were used for the visualization of the (oo)cysts. Nested PCR was applied to confirm Giardia and Cryptosporidium . Giardia and Cryptosporidium were detected in 27% and 5.5% of the 144 analyzed samples of raw sewage and effluents, respectively. A total of 33,000 cysts/L were recovered in the adhered biomass samples ( n  = 25) from different points of the aerated submerged biofilter, while 6000 oocysts/L were registered in a single point. An average of 11,800 cysts/L were found in the sludge samples ( n  = 5). The combined system exhibited a removal efficiency of Giardia cysts of 1.8 ± 1.0 log removal. The C and BIV assemblages of Giardia were identified in the raw sewage while AII was found in the treated effluent sample. It was not possible to calculate the removal efficiency of Cryptosporidium oocysts by the combined system. The combined system exhibited some potential as a suitable treatment for the removal of parasites from sewage.

Giardia is a pathogenic protozoan that has caused giardiasis outbreaks worldwide, and this water-borne disease may occur due to faults in water supply and treatment systems. To support surveillance and control programs, the monitoring of this parasite is essential in water samples. Although Giardia cyst detection is usually based on standard light microscopy, the resulting low contrasted cysts together with a wide array of particles of comparable size to the target protozoan demands a high level of observer skill and experience, as well as a long time to process the individual samples. The visualization of this protozoan can be significantly improved by using staining procedures, e.g., Lugol’s iodine in brightfield or fluorescence-based methods such as fluorescence-isothiocyanate (FITC) and 4′,6-diamidino-2-phenylindole (DAPI). However, the significant costs make fluorescence microscopy difficult to be performed in less-developed countries. Accordingly, the present study evaluated the effectiveness of inexpensive darkfield (DF) setups to acquire higher contrasted images of unstained cyst samples as a direct and rapid method for monitoring protozoa. Two low-cost custom-made DF illumination modes, transmitted and reflected, were evaluated on a standard light microscope. Examination of purified Giardia cyst suspensions with both DF setups revealed a direct correlation between morphological appearance and uptake of DAPI. Images captured under transmitted illumination showed higher contrast and sharpness when compared to the reflected images. DF microscopy might provide a simple, direct, and inexpensive method for observing Giardia cysts, which shows basic aspects of their intracellular structure, although the applicability of the method to raw water concentrates remains to be demonstrated.

Giardia and Cryptosporidium are potentially pathogenic protozoa which are ubiquitous in ambient surface water. The present study included 60 samples of surface water from three sampling sites from the Rímac River, Lima and Callao, Peru, to detect the occurrence of Giardia spp. and Cryptosporidium spp. and to perform molecular characterization of specimens found. Water samples were concentrated using the membrane filtration technique, and following elution, cysts and oocysts were visualized by direct immunofluorescence assay (IFA). For molecular characterization, tpi and bg gene fragments and 18S rRNA were amplified by nested PCR for Giardia and Cryptosporidium , respectively, followed by sequencing and phylogenetic analysis. Giardia cysts were found in 93.3% of the analyzed samples, whereas Cryptosporidium oocysts were detected in 15%. The positivity of the Giardia cysts was 86.6% ( n  = 26) in 2014, while Cryptosporidium oocysts were not detected. In 2015, both protozoa were found in raw water samples, with all 30 samples collected positive for Giardia cysts (100.0%) and 9 positive for Cryptosporidium oocysts (30.0%). Oocysts were detected in 20.0% of water samples from sites 1 (mean 5.25 oocysts/L) and 2 (mean 52.3 oocysts/L), while at site 3, oocysts were detected in 50.0% of raw water samples (mean 193.6 oocysts/L). The presence of Giardia duodenalis assemblage A was confirmed in several samples by the phylogenetic positioning of the bg and tpi genes, and the sub-assemblage AII was predominant (8/9). Sequencing for Cryptosporidium resulted in profiles compatible with Cryptosporidium hominis , Cryptosporidium meleagridis , and Cryptosporidium baileyi . This is the first time that the presence of G . duodenalis assemblage A/sub-assemblage AII and Cryptosporidium species has been reported in surface water samples in Peru. These Cryptosporidium species and the Giardia duodenalis assemblage are associated with human disease which highlights the potential risk to public health and the need to increase environmental monitoring measures to protect this water body.

Ciliated protozoa are important components of the microbial food web in various habitats, especially aquatic environments. These organisms are useful bioindicators for both environmental quality assessment and the wastewater purification process. The pathogenic parasitic protozoan species Giardia and Cryptosporidium represent a significant concern for human health, being responsible for numerous disease outbreaks worldwide. The predation of cysts and oocysts in 15 ciliate species from water and sewage samples collected in Campinas, São Paulo, Brazil were verified under laboratory conditions. The ciliated protozoan species were selected based on their mode of nutrition, and only bacterivorous and suspension-feeders were considered for the experiments. The species Blepharisma sinuosum , Euplotes aediculatus , Sterkiella cavicola , Oxytricha granulifera , Vorticella infusionum , Spirostomum minus , and Stentor coeruleus ingested cysts and oocysts, the resistance forms of Giardia spp. and Cryptosporidium spp., respectively. This is the first time that the ingestion of Giardia cysts by ciliated protozoa has been reported. These findings may contribute to a better understanding of the biological removal of these pathogens from aquatic environments.

Assessing functional diversity of communities is an efficient method to link community composition to ecosystem quality. Still, studies using functional traits of microeukaryote ciliate communities in biological wastewater treatment plants are lacking. The present work explores the functional diversity of the ciliate protist community in a wastewater treatment plant (WWTP) operating with a combined UASB-activated sludge system, and specifically to: 1) investigate the taxonomic and functional composition of the ciliate communities over time; 2) compare taxonomic and functional diversity indices with regard to its applicability in WWPS; 3) assess the relationship between the ciliate community’s functional composition and the WWTPs temporal conditions; and 4) investigate the potential use of functional diversity as an indicator of WWTP efficiency. Totally, we recorded 21 ciliate species throughout 37 samplings. The number of species was low compared to other plants. Bacterivorous and flake-forming species were the main functional strategies found in the samples. The correlation between taxonomic and functional richness was significant, indicating a functionally redundant community. There was a correlation between the Simpson and Rao’s quadratic entropy indexes suggesting that loss of taxonomic diversity leads to a loss of functional diversity. The homogeneity of the measured physical and chemical data led to functional homogenization and redundancy (homogenous CWM) of the ciliate community. The functional diversity is positively correlated with parameters of removal efficiency, indicating a promising application in WWTPs. Future studies will broaden knowledge on functional diversity in biological wastewater treatment systems, this being a first step with the unprecedented application of this methodology in artificial ecosystems.

After a gastroenteritis outbreak of unknown etiology in the municipality of Sebastião da Grama, São Paulo, Brazil, we conducted a parasitological survey to establish the epidemiological profile of enteroparasitosis in children and staff members attending the public urban schools in operation in town. The cross-sectional study evaluated 172 children aged 11 months to 6 years old and 33 staff members aged 19 to 58 years old. Overall, 96 (55.81%) children and 20 (60.61%) staff members were mono-parasitized, while 58 (33.72%) children and 4 (12.12%) workers were poly-parasitized. Protozoa (88.37%; 72.73%) was more prevalent than helminthes (3.48%; 0%) in children and staff members respectively.Blastocystis spp. was the most prevalent parasite in children (86.63%) and staff members (66.67%). The age of 1 year old or less was found to be associated with increased prevalence of giardiasis [OR = 13.04; 95%CI 2.89-58.91; p = 0.00] and public garbage collection was identified as a protective factor against intestinal helminth infections [OR = 0.06; 95%CI 0.00-0.79; p = 0.03]. Although most of the children tested positive for Blastocystis spp. and also presented clinical signs/symptoms (62.2%), this association was not statistically significant [OR = 1.35; 95%CI 0.53-3.44; p = 0.51]. Intestinal parasites still represent a public health concern and this study underscores the importance of further investigations to better understand the pathogenic role of Blastocystis spp.

The aims of the present study were (1) to evaluate the abundance and taxonomic composition of ciliated protozoa in the activated sludge of a full-scale combined anaerobic-aerobic system operating in a tropical country and (2) to study the relationship between the effluent quality, the physicochemical variables, and the ciliates present in the operating system. The total ciliate fauna of the activated sludge of the Piçarrão Wastewater Treatment Plant (Piçarrão WWTP) was composed of 36 morphospecies belonging to 33 genera. These included 21 species observed in the activated sludge samples on the day of collection and 15 species found in cultures. The activated sludge of the Piçarrão WWTP contained a diversified ciliate community composed mainly of indicator organisms. The most frequently occurring morphospecies were Aspidisca cicada , Vorticella spp., Gastronauta aloisi , Acineria uncinata , and Epistylis plicatilis complex. These results showed that satisfactory operating conditions prevailed at the Piçarrão WWTP. In the combined UASB–activated sludge system, the presence of Aspidisca cicada suggests the occurrence of denitrification in the process while the presence of Acineria uncinata and G. alosi indicates the removal of carbonaceous organic matter.