Epidemic and endemic seroprevalence of antibodies to Cryptosporidium and Giardia in residents of three communities with different drinking water supplies.
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This study was carried out to compare cryptosporidiosis and giardiasis seroprevalence rates in residents of three communities. Community (Com 1) uses drinking water from deep wells, community 2 (Com 2) uses surface water from a protected watershed, and community 3 (Com 3) uses surface water frequently containing Cryptosporidium oocysts and Giardia cysts. Unfiltered drinking water from each community was collected at the tap and tested for Cryptosporidium oocysts and Giardia cysts during the 12 months in which sera were collected for testing. No oocysts or cysts were detected in the water from the Com 1 deep wells; oocysts and cysts were detected intermittently in the drinking water from the other two communities. A waterborne outbreak of cryptosporidiosis occurred in a municipality adjacent to Com 3 six months into this 12-month study. Sera from residents of each of the communities were collected proportionately by month and by population size. Coded sera were tested for IgG to Cryptosporidium using a previously developed Western blotting method. The presence or absence of bands at 15-17 kD and/or 27 kD was recorded for the 1,944 sera tested. Definite bands at 15-17 kD and/or 27 kD were detected in 981 (50.5%) of the sera. A total of 33.2% of sera from Com 1 (community using deep wells) were positive using the same criteria compared with 53.5% (Com 2) and 52.5% (Com 3) of sera from the two communities using surface drinking water. Both bands (15-17 kD plus 27 kD) were detected in 582 sera (29.9%) from the three communities: 14.1% of sera from Com 1 compared with 32.7% from Com 2 and 31.5% from Com 3. These findings are consistent with a lower risk of exposure to Cryptosporidium from drinking water obtained from deep well sources. However, analysis of results by calendar quarter showed a significant (P < 0.001) increase in the number of Com 3 positive sera (compared with Com 1) following the waterborne outbreak. Without this outbreak-related observation, a significant overall difference in seropositivity would not have been seen. We also observed that in sera from the community affected by the outbreak, the presence on immunoblots of both Cryptosporidium bands appeared to be the best indicator of recent infection. Seroprevalence rates using an ELISA to detect IgG to Giardia were estimated using the same sera. Overall 30.3% (590 of 1,944) of sera were positive by the ELISA. A total of 19.1% of sera from Com 1, 34.7% from Com 2 and 16.0% from Com 3 were seropositive. Rates for both Com 3 and Com 1 did not change significantly over time. In Com 2, rates decreased significantly (P < 0.001) during the last half of the study period (third and fourth calendar quarters). The reasons for the decrease in seroprevalence in Com 2 sera are presently not known. These studies show intriguing associations between seroprevalence, outbreak-related laboratory serologic data, and patterns of parasite contamination of drinking water. Further studies are required to validate the serologic approach to risk assessment of waterborne parasitic infections at a community level. (+info)
Influence of refrigeration and formalin on the floatability of Giardia duodenalis cysts.
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Giardia duodenalis cysts obtained from fresh fecal samples, fecal samples kept under refrigeration and fecal samples treated with formalin were studied as to their floatability on sucrose solutions with the following specific gravities: 1,040 kg/m3; 1,050 kg/m3; 1, 060 kg/m3; 1,070 kg/m3; 1,080 kg/m3; 1,090 kg/m3; 1,100 kgm3; 1,150 kg/m3; 1,200 kg/m3; and 1,250 kg/m3, contained within counting-chambers 0.17 mm high. Cysts that floated on and those settled down as sediments were counted, and had their percentages estimated. Sucrose solutions of 1,200 kg/m3 specific gravity (the average specific gravity of diluting liquids employed in floatation techniques) caused to float 77.7%, 78.4% and 6.6% of the G. duodenalis cysts obtained, respectively, from fresh fecal samples, fecal samples kept under refrigeration, and fecal samples treated with formalin. Cysts obtained both from fresh fecal samples and fecal samples kept under refrigeration presented similar results concerning floatability. It was observed, however, that the treatment of feces with formalin diminished the cysts floatability under the various specific gravities studied. This results should influence, the recommendations for transport and storage of fecal samples used for parasitological coproscopy. (+info)
Molecular systematics of the parasitic protozoan Giardia intestinalis.
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The long-standing controversy regarding whether Giardia intestinalis is a single species prevalent in both human and animal hosts or a species complex consisting of morphologically similar organisms that differ in host range and other biotypic characteristics is an issue with important medical, veterinary, and environmental management implications. In the past decade, highly distinct genotypes (some apparently confined to particular host groups) have been identified by genetic analysis of samples isolated from different host species. The aim of this study was to undertake a phylogenetic analysis of G. intestinalis that were representative of all known major genetic groups and compare them with other Giardia species, viz. G. ardeae, G. muris, and G. microti. Segments from four "housekeeping" genes (specifying glutamate dehydrogenase, triose phosphate isomerase, elongation factor 1 alpha, and 18S ribosomal RNA) were examined by analysis of 0.48-0.69-kb nucleotide sequences determined from DNA amplified in polymerase chain reactions from each locus. In addition, isolates were compared by allozymic analysis of electrophoretic data obtained for 21 enzymes representing 23 gene loci. The results obtained from these independent techniques and different loci were essentially congruous. Analyses using G. ardeae and/or G. muris as outgroups supported the monophyly of G. intestinalis and also showed that this species includes genotypes that represent at least seven deeply rooted lineages, herein designated assemblages A-G. Inclusion of G. microti in the analysis of 18S rRNA sequence data demonstrated the monophyly of Giardia with the same median body morphology but did not support the monophyly of G. intestinalis, instead placing G. microti within G. intestinalis. The findings support the hypothesis that G. intestinalis is a species complex and suggest that G. microti is a member of this complex. (+info)
Immune response to Giardia duodenalis.
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The intestinal protozoan Giardia duodenalis is a widespread opportunistic parasite of humans and animals. This parasite inhabits the upper part of the small intestine and has a direct life cycle. After ingestion of cysts, which are the infective stage, the trophozoites emerge from the cysts in the duodenum and attach to the small intestinal mucosa of the host. Since the migration of trophozoites from the lumen of the intestine into surrounding tissues is an unusual occurrence, the immune response to Giardia remains localized. The identification of antigens that play a role in acquired immunity has been difficult because of the occurrence of antigenic variation and because, Giardia being an ubiquitous organism, it is possible that the antigenic profiles of isolates from different geographic areas will vary. Innate-immunity mechanisms play a role in the control and/or severity of the infection. Both humoral and cell-mediated immune responses play a role in acquired immunity, but the mechanisms involved are unknown. A variety of serological assays have been used to detect circulating antibodies in serum. Because of the biological characteristics of the parasite and the lack of suitable antigens, the sensitivity of serological assays remains poor. On the other hand, detection of antigens in feces of infected patients has met with success. Commercial kits are available, and they are reported to be more sensitive than microscopic examination for the detection of giardiasis on a single specimen. (+info)
Influence of bacteria from the duodenal microbiota of patients with symptomatic giardiasis on the pathogenicity of Giardia duodenalis in gnotoxenic mice.
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Recent studies have shown that the intestinal microbiota is essential for the pathogenicity but not for the multiplication of Giardia duodenalis in the intestinal lumen. The microbial components responsible for this phenomenon are not known. Twenty-eight facultative and three strictly anaerobic micro-organisms were isolated from the dominant duodenal microbiota of five patients with symptomatic giardiasis. The bacterial combinations from each patient were associated with groups (GN) of germ-free mice. Five days after the association, when their faecal populations ranged from 10(7) to 10(9) cfu/g, all groups were inoculated intragastrically with 10(5) viable trophozoites of G. duodenalis strain BT6. Two groups of germ-free (GF) and conventional (CV1) mice were also infected. Gnotobiotic animals were killed 10 days after infection and GF and CV1 animals were killed 10, 20 and 30 days after infection. More marked pathological alterations were detected in CV1 mice when compared with GF animals. Gnotobiotic animals showed intermediate pathological alterations between CV1 and GF mice. The CV1 and GF groups became infected by day 3 and faecal cyst levels were similar in both groups throughout the experiment. Total and G. duodenalis-specific IgA levels in the intestinal fluid and G. duodenalis-specific IgM and IgG levels in the serum increased during the infection and were higher in CV1 animals at all times tested when compared with GF mice. The present results confirm the stimulatory activity of the intestinal microbiota on the pathogenicity of G. duodenalis, and some combinations of microbial components of the dominant duodenal ecosystem from patients with symptomatic giardiasis can partially develop this function. However, none of these combinations was able to stimulate the protozoan pathogenicity in the same manner as the entire intestinal microbiota. (+info)
Effect of sample holding time on recovery of Cryptosporidium oocysts and Giardia cysts from water samples.
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U.S. Environmental Protection Agency methods for analysis of water for Cryptosporidium and Giardia stipulate maximum sample holding times which are not always practical to comply with. A spiking experiment indicated that holding times of up to 2 weeks had no significant effect on recovery of these parasites from 10-liter samples of raw water in plastic carboys. (+info)
Stage-specific expression and targeting of cyst wall protein-green fluorescent protein chimeras in Giardia.
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In preparation for being shed into the environment as infectious cysts, trophozoites of Giardia spp. synthesize and deposit large amounts of extracellular matrix into a resistant extracellular cyst wall. Functional aspects of this developmentally regulated process were investigated by expressing a series of chimeric cyst wall protein 1 (CWP1)-green fluorescent protein (GFP) reporter proteins. It was demonstrated that a short 110 bp 5' flanking region of the CWP1 gene harbors all necessary cis-DNA elements for strictly encystation-specific expression of a reporter during in vitro encystation, whereas sequences in the 3' flanking region are involved in modulation of steady-state levels of its mRNA during encystation. Encysting Giardia expressing CWP1-GFP chimeras showed formation and maturation of labeled dense granule-like vesicles and subsequent incorporation of GFP-tagged protein into the cyst wall, dependent on which domains of CWP1 were included. The N-terminal domain of CWP1 was required for targeting GFP to regulated compartments of the secretory apparatus, whereas a central domain containing leucine-rich repeats mediated association of the chimera with the extracellular cyst wall. We show that analysis of protein transport using GFP-tagged molecules is feasible in an anaerobic organism and provides a useful tool for investigating the organization of primitive eukaryotic vesicular transport. (+info)
Core histones of the amitochondriate protist, Giardia lamblia.
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Genes coding for the core histones H2a, H2b, H3, and H4 of Giardia lamblia were sequenced. A conserved organism- and gene-specific element, GRGCGCAGATTTVGG, was found upstream of the coding region in all core histone genes. The derived amino acid sequences of all four histones were similar to their homologs in other eukaryotes, although they were among the most divergent members of this protein family. Comparative protein structure modeling combined with energy evaluation of the resulting models indicated that the G. lamblia core histones individually and together can assume the same three-dimensional structures that were established by X-ray crystallography for Xenopus laevis histones and the nucleosome core particle. Since G. lamblia represents one of the earliest-diverging eukaryotes in many different molecular trees, the structure of its histones is potentially of relevance to understanding histone evolution. The G. lamblia proteins do not represent an intermediate stage between archaeal and eukaryotic histones. (+info)