Regina Maura Bueno Franco

Occurrence of Cryptosporidium oocysts and Giardia cysts in raw water from the Atibaia River, Campinas, Brazil.

Cryptosporidium parvum and Giardia duodenalis are waterborne parasites that have caused several outbreaks of gastrointestinal disease associated with drinking water. Due to the lack of studies about the occurrence of these protozoa in water in the Southeast of Brazil, an investigation was conducted to verify the presence of cysts and oocysts in superficial raw water of the Atibaia River. The water samples were submitted to membrane filtration (3.0 microm) and elution was processed by (1) scraping and rinsing of membrane (RM method) and (2) acetone-dissolution (ADM method). Microbiologic and chemical parameters were analyzed. Aliquots of the pellets were examined by immunofluorescence (Merifluor, Meridian Diagnostics, Cincinnati, Ohio). All water samples were positive for Cryptosporidium and Giardia, in spite of the high turbidity. Higher recovery rates occurred in samples treated by the RM method than by the ADM technique. The goal for future work is the assessment of viability of cysts and oocysts to determine the public health significance of this finding.

Transmission of toxoplasmosis (Toxoplasma gondii) by foods.

Protozoan foodborne diseases are generally underrecognized. Toxoplasma gondii is the causative agent of toxoplasmosis, one of the most prevalent parasitic infections to humans and domestic animals. The most likely source of T. gondii occurring through food is the consumption of raw or undercooked meat contaminated with tissue cysts. Sporulated T. gondii oocysts, from the feces of infected cats, present in the environment are a potential source of infection. The ingestion of water contaminated with oocysts and the eating of unwashed raw vegetables or fruits were identified as an important risk factor in most epidemiological studies. This review presents information and data to show the importance of T. gondii transmission by foods.

Evaluation of tropical water sources and mollusks in southern Brazil using microbiological, biochemical, and chemical parameters

Florianópolis, a city located in the Santa Catarina State in southern Brazil, is the national leading producer of bivalve mollusks. The quality of bivalve mollusks is closely related to the sanitary conditions of surrounding waters where they are cultivated. Presently, cultivation areas receive large amounts of effluents derived mainly from treated and non-treated domestic, rural, and urban sewage. This contributes to the contamination of mollusks with trace metals, pesticides, other organic compounds, and human pathogens such as viruses, bacteria, and protozoan. The aim of this study was to perform a thorough diagnosis of the shellfish growing areas in Florianópolis, on the coast of Santa Catarina. The contamination levels of seawater, sediments, and oysters were evaluated for their microbiological, biochemical, and chemical parameters at five sea sites in Florianópolis, namely three regular oyster cultivation areas (Sites 1, 2, and oyster supplier), a polluted site (Site 3), and a heavily polluted site (Site 4). Samples were evaluated at day zero and after 14 days. Seawater and sediment samples were collected just once, at the end of the experiment. Antioxidant defenses, which may occur in contaminated environments in response to the increased production of reactive oxygen species (ROS) by organisms, were analyzed in oysters, as well as organic compounds (in oysters and sediment samples) and microbiological contamination (in oysters and seawater samples). The results showed the presence of the following contaminants: fecal coliforms in seawater samples (four sites), human adenovirus (all sites), human noroviruses GI and GII (two sites), Hepatitis A viruses (one site), JC Polyomavirus in an oyster sample from the oyster supplier, Giardia duodenalis cysts, and Cryptosporidium sp oocysts (one site). Among organochlorine pesticides, only DDT (dichlorodiphenyltrichloroethane) and HCH (hexachlorocyclohexane) were detected in some sediment and oysters samples in very low levels; site 4 had the highest concentrations of total aliphatic hydrocarbons, PAHs, and linear alkylbenzenes (LABs) found either in oysters or in sediment samples. The major concentration of fecal sterol coprostanol was found at site 4, followed by site 3. After 14 days of allocation in the four selected sites, there was a significant difference in the enzymes analyzed at the monitored spots. The detection of different contaminants in oysters, seawater, and sediment samples in the present study shows the impact untreated or inadequately treated effluents have on coastal areas. These results highlight the need for public investment in adequate wastewater treatment and adequate treatment of oysters, ensuring safe areas for shellfish production as well as healthier bivalve mollusks for consumption.

Transmission of Chagas Disease (American Trypanosomiasis) by Food

In April 2009, the centenary of the discovery of the American trypanosomiasis, or Chagas disease, was celebrated. A hundred years after the discovery, little has been invested in diagnostics and treatment because the disease affects mainly poor people in developing countries. However, some changes in the epidemiology of the disease are of great importance today. Chagas disease transmitted through food is a public health concern in all areas where there is a reservoir of Trypanosoma cruzi in wild animals (e.g., mammals and marsupials) and/or where infected triatomine bugs are in contact with human food source items (especially fruits and vegetables). Recently, several outbreaks of illness related to the ingestion of food contaminated with T. cruzi have been recorded in Brazil, Colombia, and Venezuela.

Molecular characterization of intestinal protozoa in two poor communities in the State of São Paulo, Brazil.

BackgroundSeveral 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.MethodsMultiple 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.ResultsBy 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.ConclusionsThis 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.

Giardiose e criptosporidiose em creches no município de Campinas, SP

The prevalence of the Giardia duodenalis and Cryptosporidium parvum infections were studied in children 2-60 months old (n = 310) grouped to sex and age, enrolled in 8 day-care centers. Giardiosis was found in 42 (13.5%) of the children and, was most frequent in the age group corresponding to 19- 24 months old although children at the first six months of life also presented positive faecal samples. Statistical analysis indicated no association between sex ip > 0,05) and prevalence for Giardia infection. C. parvum was detected in 20 (6.4%) of the children. Children seven to twelve months old showed the greatest prevalence of this parasite. There were not oocysts in the stools from the children 25-30 months and from those older than 36 months. Oocysts were detected in 4 children from the 0-6 months old group. There was no significative difference on sex (p = 0,09) and day-care facility (p = 0,068) for C. parvum infection while age was associated with infection (p = 0,004). Epidemiological aspects were studied by familiar inquiry, by parasitological examinations of stools from the contact- members of each case and from animals present at the residence. Prevalence for C. parvum was higher when at least one of the parents was a professional in the Human Health Centers (73.6% positives), when there was report of concomitant disease (52.6%positives) and when diarrhoea was the main symptom at the collection of the faecal specimens (78.9% positives). Faecal examinations of the domestic animals were negatives for these two parasites.

Cryptosporidium spp. and Giardia spp. in surface water supply of Campinas, Southeast Brazil

Surface water contaminated by domestic sewage discharges is a potential source of pathogens, including protozoa. During 2005-2006, the source water (Atibaia River) of the Surface Water Treatment Plant (WTP) of Campinas city, São Paulo, Brazil was sampled to obtain an assessment of Cryptosporidium oocyst and Giardia cyst concentrations. Calcium carbonate flocculation (CCF) and membrane filtration (MF) concentration techniques, with and without purification by immunomagnetic separation (IMS) were evaluated. The cysts and oocysts were detected by immunofluorescence assay (IFA) and confirmed by differential interference contrast (DIC). Membrane filtration method generally produced higher recovery efficiency. Giardia spp. was detected in 87.5% of the water samples analyzed with densities ranging from 2.5 to 120 cysts per L. Cryptosporidium spp were detected in 62.5% and the concentrations ranged from 15 to 60 oocysts per L. Cryptosporidium oocyst and Giardia cyst concentrations detected in this study were elevated and are associated with discharge of untreated sewage in Atibaia River. Measures should be taken to protect surface water from sources of contamination.

Occurrence of Giardia cysts and Cryptosporidium oocysts in activated sludge samples in Campinas, SP, Brazil

Giardia and Cryptosporidium have caused several outbreaks of gastroenteritis in humans associated with drinking water. Contaminated sewage effluents are recognized as a potential source of waterborne protozoa. Due to the lack of studies about the occurrence of these parasites in sewage samples in Brazil, we compared the efficiency of two procedures for concentrating cysts and oocysts in activated sludge samples of one sewage treatment plant. For this, the samples were submitted to i) concentration by the ether clarification procedure (ECP) and to ii) purification by sucrose flotation method (SFM) and aliquots of the pellets were examined by immunofluorescence. Giardia cysts were present in all samples (100.0%; n = 8) when using ECP and kit 1 reagents, while kit 2 resulted in six positive samples (85.7%; n = 7). As for SFM, cysts were detected in 75.0% and 100.0% of these samples (for kit 1 and 2, respectively). Regarding Cryptosporidium, two samples (25.0%; kit 1 and 28.5% for kit 2) were detected positive by using ECP, while for SFM, only one sample (examined by kit 1) was positive (12.5%). The results of the control trial revealed Giardia and Cryptosporidium recovery efficiency rates for ECP of 54.5% and 9.6%, while SFM was 10.5% and 3.2%, respectively. Considering the high concentration detected, a previous evaluation of the activated sludge before its application in agriculture is recommended and with some improvement, ECP would be an appropriate simple technique for protozoa detection in sewage samples.

Occurrence of Cryptosporidial Oocysts and Giardia Cysts in Bottled Mineral Water Commercialized in the City of Campinas, State of São Paulo, Brazil

The consumption of bottled mineral water has significantly increased in Brazil so that it is in the interest of public health to determine the parasitological and microbiological status of some brands of Brazilian mineral water available in the town of Campinas, São Paulo, Brazil. For this purpose, detection of protozoa by direct immunofluorescence technique and microbiological parameters were determined for each specimen after membrane filtration. Giardia cysts were not present while cryptosporidial oocysts were detected in two samples. The counts of protozoa varied from 0.2 to 0.5 oocysts/l. The detected level of Pseudomonas aeruginosa and heterotrophic bacteria reflected the level of organic enrichment of the water.

First report of Cryptosporidium spp. oocysts in oysters (Crassostrea rhizophorae) and cockles (Tivela mactroides) in Brazil.

The consumption of oysters and cockles, which are usually eaten raw or lightly-cooked, can cause outbreaks of human diseases, especially if these shellfish are harvested from polluted areas. In Brazil data about the occurrence of pathogens, like hepatitis A virus, in shellfish have been reported but research on natural contamination for pathogenic protozoa is still non-existent. Cryptosporidium oocyst contamination of oysters (Crassostrea rhizophorae) and cockles (Tivela mactroides) was evaluated during two different periods in a coastal area from São Paulo, Brazil. From June to November 2005, and from July to December 2006, 180 mollusks were harvested for tissue examination. The gills and gastrointestinal tract (n = 36 pools) were carefully extracted from the animals and homogenized in a tissue homogenizer by adding surfactant Tween 80 (0.1%). Immunofluorescence assays were performed and Cryptosporidium oocysts were detected in 50.0% of gill pools of cockles and 10.0% of gill pools of oysters. In order to evaluate seawater quality in shellfish growing areas, total levels of thermotolerant coliforms, Escherichia coli and enterococci were determined. This is the first time that Cryptosporidium oocysts were found in shellfish from the coastal region of Brazil, and to the best of our knowledge it is also the first report in Latin America and the case might be of public health importance, reflecting the extension of the contamination on seafood, requiring a need for quality control standards.